Meiosis regulating compounds

ABSTRACT

Certain novel sterol derivatives can be used for regulating the meiosis in oocytes and in male germ cells.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No. 09/332,235 filed on Jun. 14, 1999 now abandoned, which is a continuation of PCT/DK99/00263 filed May 11, 1999 and claims priority under 35 U.S.C. 119 of Danish applications PA 1998 00657 filed May 13, 1998 and PA 1998 00811 filed on Jun. 19, 1998, European application 98250166.0 filed May 14, 1998, and U.S. provisional applications No. 60/086,306 filed on May 21, 1998 and No. 60/092,983 filed on Jul. 16, 1998, the contents of which are fully incorporated herein by reference.

FIELD OF THIS INVENTION

The present invention relates to certain pharmacologically active compounds, to pharmaceutical composition containing certain compounds as active substance and to their use as medicaments. More particularly, it has been found that the derivative described herein can be used for regulating the meiosis.

BACKGROUND OF THIS INVENTION

Meiosis is the unique and ultimate event of germ cells on which sexual reproduction is based. Meiosis comprises two meiotic divisions. During the first division, exchange between maternal and paternal genes take place before the pairs of chromosomes are separated into the two daughter cells. These contain only half the number (1n) of chromosomes and 2c DNA. The second meiotic division proceeds without a DNA synthesis. This division, therefore, results in the formation of the haploid germ cells with only 1c DNA.

The meiotic events are similar in the male and female germ cells, but the time schedule and the differentiation processes which lead to ova and to spermatozoa differ profoundly. All female germ cells enter the prophase of the first meiotic division early in life, often before birth, but all are arrested as oocytes later in the prophase (dictyate state) until ovulation after puberty. Thus, from early life the female has a stock of oocytes which is drawn upon until the stock is exhausted. Meiosis in females is not completed until after fertilisation, and results in only one ovum and two abortive polar bodies per germ cell. In contrast, only some of the male germ cells enter meiosis from puberty and leave a stem population of germ cells throughout life. Once initiated, meiosis in the male cell proceeds without significant delay and produces 4 spermatozoa.

Only little is known about the mechanisms which control the initiation of meiosis in the male and in the female. In the occyte, new studies indicate that follicular purines, hypoxanthine or adenosine, could be responsible for meiotic arrest (Downs, S. M. et al. Dev. Biol. 82 (1985) 454-458; Eppig, J. J. et al. Dev. Biol. 119 (1986) 313-321; and Downs, S. M. Mol. Reprod. Dev. 35 (1993), 82-94). The presence of a diffusible meiosis regulating substance was first described by Byskov et al. in a culture system of fetal mouse gonads (Byskov, A. G. et al. in Dev. Biol. 52 (1976), 193-200). A meiosis activating substance, (MAS) was secreted by the fetal mouse ovary in which meiosis was ongoing, and a meiosis preventing substance (MPS) was released from the morphologically differentiated testis with resting, non-meiotic germ cells. It was suggested that the relative concentrations of MAS and MPS regulated the beginning, arrest and resumption of meiosis in the male and in the female germ cells (Byskov, A. G. et al. in The Physiology of Reproduction (eds. Knobil, E. and Neill, J. D., Raven Press, New York (1994)). Clearly, if meiosis can be regulated, reproduction can be controlled. A recent article (Byskov, A. G. et al. in Nature 374 (1995), 559-562) describes the isolation from bull testes and from human follicular fluid of certain sterols that activate oocyte meiosis. Unfortunately, these sterols are rather labile and utilisation of the interesting finding would thus be greatly facilitated if more stable meiosis activating compounds were available.

In Aust. J. Chem. 35 (1982), 629-640, Horn et al. deals with compounds possibly having biological activity (insect moulting hormones). Examples of compounds specifically mentioned therein are 5-cyano-5βcholest-7-en-3-one; 5-cyano-5βcholest-7-en-3β-ol; 5-methyl-5β-cholest-7-en-3-one; 5-methyl-5β-cholest-7-en-3α-ol; and 5-methyl-5β-cholest-7-en-3β-ol.

In Bull. Soc. Chim. Fr. (1971), 2037-2047, Levisalles et al., cholesta-4,8(14)-dien-3-one is described as an intermediate.

In Just. Lieb. Ann. Chem. 542 (1939), 218-224, Windaus et al. mentions cholesta-4,7-dien-3-one; cholesta-4,7-dien-3α-ol; and cholesta-4,7-dien-3β-ol as intermediates.

In Pharm. Bull. 1 (1953), 224-227, Arima mentions cholesta-4,8-dien-3-one as intermediate.

In Lipids 13 (1978), 704 et seq., Kandutsch et al. describes some cholestane derivatives which may be potent inhibitors of sterol synthesis. Compounds specifically mentioned therein are, in FIG. 1, 3β,7α-dihydroxycholest-5-ene; 3β,7β-dihydroxycholest-5-ene; 3β-hydroxycholest-5-en-7-one; 3β-hydroxycholest-7-one; 7α-hydroxycholest-4-en-3-one; in FIG. 2 (compounds 1-5), cholest-3,6-dione; 3β-hydroxycholest-6-one; 3β,6β-dihydroxycholestane; cholest-4-en-3,6-dione; 3β,5α,6β-trihydroxycholestane; in FIG. 3, 3β,5α-dihydroxycholestane; 3β,4β-dihydroxycholest-5-ene; and, in FIG. 4 (compounds 1, 3, 4 and 5), cholest-2-en-6-one; cholest-4,6-dien-3-one; cholest-4,7-dien-3-one; cholest-3,5-dien-7-one.

Danish patent application with publication number 130,992, deals with compounds possibly having progestomimetic properties. Examples of compounds specifically mentioned therein are 19-nor-21-methylpregna-4,9-dien-17α-hydroxy-3,20-dione; 19-nor-21-methyl-pregna-4,9-dien-17α-acetoxy-3,20-dione; 19-nor-21,21-dimethylpregna-4,9-diene-17α-hydroxy-3,20-dione; and 17α-21,21-dimethyl-19-nor-pregna-4,9-dien-3,20-dione.

In the Danish patent application with publication number 136,909, 3α-acetoxy-24-nor-cholan-23-one; 3α-hydroxy-26,27-di-nor-23-trans-5β-cholest-23-en-25-carboxylic acid methyl ester; 3α-hydroxy-26,27-di-nor-5β-cholesta-25-carboxylic acid methyl ester; 3-keto-26,27-di-nor-5β-cholesta-25-carboxylic acid methyl ester; 3-keto-4-bromo-26,27-di-nor-5β-cholesta-25-carboxylic acid methyl ester; 3-keto-26,27-di-nor-cholest4-en-25-carboxylic acid methyl ester; 3β-acetoxy-26,27-di-nor-cholesta-3,5-dien-25-carboxylic acid methyl ester; 3β-hydroxy-26,27-di-nor-cholest-5-en-25-carboxylic acid methyl ester; 3-keto-26,27-di-nor-cholest-4,6-dien-25-carboxylic acid methyl ester; 3β-hydroxy-26,27-di-nor-cholesta-3,5,7-trien-25-carboxylic acid methyl ester; and 3β-hydroxy-26,27-di-nor-cholesta-5,7-dien-25-carboxylic acid methyl ester are mentioned as intermediates.

Danish patent application with publication number 146,390, deals with compounds possibly having pharmacological properties, e.g. an inhibiting action on the production of serum cholesterol. Examples of compounds specifically mentioned therein are 3β,22-di-acetoxycholesta-5-en-25-ol; 3β,22-diacetoxy-25-fluorocholesta-5-ene; 22-hydroxycholesta-5-en-25-fluoro-3β-hemisuccinate; 3β,22-diacetoxy-25-dichlorocholesta-5-ene; 3β,22-dihydroxy-25-chlorocholesta-5-ene; 22-hydroxy-25-chlorocholesta-5-en-3β-hemisuccinate; 3β,22-dihydroxy-25-bromocholesta-5-ene; and 3β,22-dihydroxy-25-fluorocholesta-5-ene.

In the Danish patent applications with publication numbers 156,726 and 156,644, cholenic acid; 3β-acetoxycholesta-5-en-25-des-dimethyl-24-one; 3,24-diacetoxycholesta-25-des-dimethyl-5,23-diene; 3β-acetoxycholesta-25-des-methyl-5-en-24-difluoro-25-one; and 3β-acetoxy-24-difluorocholesta-5,7-dien-25-ol are mentioned as intermediates.

In the Danish patent application with publication number 158,790, 3β-hydroxy-cholest-5-en-24-one; 3β-acetoxycholest-5-en-24-one; 5β-cholest-24-one; 5β-cholestan-24α-homo-24-one; 3α,6α-dihydroxy-5β-cholest-24-one; 3α,6α-diacetoxy-5β-cholest-24-one; 3α,6α-diacetoxy-5β-cholest-24α-homo-24-one; 3α,6α-dihydroxy-5β-cholest-24α,24β-bis-homo-24-one; 3α-hydroxy-5β-cholest-24-one; 3α-acetoxy 5β-cholesta-24-one; 3α-benzoyl-oxy-5β-cholesta-24-one; 3α-ethyloxycarbonyloxy-5β-cholesta-24-one; 3α-hydroxy-5β-cholestan-24α-homo-24-one; 3α-hydroxy-24α,24β-bis-homo-5β-cholestane; 3β-hydroxy-cholesta-5,7-dien-24-one; 3β-acetoxycholesta-5,7-dien-24-one; 1α,3β-dihydroxycholesta-5,7-diene-24-one; and 1α,3β-diacetoxycholesta-5,7-diene-24-one are mentioned as intermediates.

Danish patent application with publication number 159,456, deals with compounds possibly having utility in the treatment of gall diskinese. Examples of compounds specifically mentioned therein are chenodeoxycholic acid; ursodeoxycholic acid; trimebutyn salt of chenodeoxycholic acid; and trimebutyn salt of ursodeoxycholic acid.

In the Danish patent application with publication number 162,648, 3β,25-dihydroxy-cholest-5-en-24-one; 3β-acetoxycholest-5-en-24-one; 3β-acetoxy-25-hydroxycholest-5-en-24-one; 3β,25-dihydroxycholest-5-en-24-one; 3β-hydroxycholest-5-en-24-one; 3β-hydroxy-25-hydroperoxycholest-5-en-24-one; 3β,24,25-trihydroxycholest-5-ene; 1α,3β-dihydroxycholest-5-en-24-one; 1α,3β,25-trihydroxycholest-5-en-24-one; 1α,3β,24,25-tetrahydroxycholest-5-en-24-one are mentioned as intermediates.

In the Danish patent application with publication number 165,410, 3α-acetoxy-7α-bromocholest-5-ene; 3α-acetoxycholesta-5,7-diene; and 3α-acetoxy-25-hydroxycholesta-5,7-diene are mentioned as intermediates.

In the Danish patent application with publication number 165,695, 3β,25-dihydroxy-26,27-hexafluorocholest-5-ene; and 1α,3β,25-trihydroxy-26,27-hexafluorocholest-5-ene are mentioned as intermediates for the preparation of vitamin D analogues.

Danish patent application with publication number 167,220 deals with compounds possibly having utility for the treatment of liver disorders. An examples of a compound specifically mentioned therein is 3α,7α,12α,24R,26,27-hexahydroxycholestane.

In U.S. Pat. No. 4,425,274, the compounds 3α-hydroxy-7-cholanic acid; 3α,7α-dihydroxycholanic acid; 3α,7β-dihydroxycholanic acid; and lithium 3α,7β, dihydroxycholanate are described as intgermediates.

In the Norwegian patent application with publication number 144,264, cholesta-1,4,-6-trien-3-one; cholest-5-en-1α,3β-diol; 1α-hydroxycholesta-4,6-diene-3-one; 1α,3β, dihydroxycholest-5-ene; 25-hydroxycholesta-1,4,6-triene-3-one; and 1α,3β-25-trihydroxycholest-5-ene are mentioned as intermediates for the preparation of 1α-hydroxy steroids of the cholestane serie.

Norwegian patent application with publication number 158,423, deals with compounds possibly having utility in the treatment of biliary dyskinesis. An example of a compound specifically mentioned therein is 3α,7β-dihydroxydeoxycholic acid.

In the Norwegian patent application with publication number 162,562, 3α,7α-dihydroxydeoxycholic acid; 7-ketodeoxycholic acid; 3α,7β-dihydroxydeoxycholic acid; cholic acid; 7-ketocholic acid; 3α,7β,12α-cholic acid; and 12-ketocholic acid are mentioned as being intermediates in the preparation of ursodeoxycholanic acid.

In the Norwegian patent application with publication number 162,665, cholic acid; 3α-hydroxy-7-ketocholic acid; 3α,7α-diacetoxycholic acid; 3α,7α-diacetoxy-12-ketocholic acid; 3α,7α-dihydroxydeoxycholic acid; 7-ketodeoxycholic acid; 3α,7β-dihydroxydeoxycholic acid; and 3α,7β-dihydroxy-12-ketocholic acid are mentioned as being intermediates in the preparation of ursodeoxycholanic acid.

In the Norwegian patent application with publication number 303,450, cholic acid; cholic acid methyl ester; 3-acetyicholic acid methyl ester; 3-(2-propenyl)cholic acid methyl ester; 3-(3-hydroxypropyl)cholic acid-3-methyl ester; desoxycholic acid; 12-keto-desoxycholic acid; 3β-acetyloxy-12-keto-desoxycholic acid; 3β-(hydroxyethyloxy)cholic acid methyl ester; 3β-(hydroxypropyloxy)cholic acid methyl ester; 3β-(hydroxybutyloxy)cholic acid methyl ester; 3β-(hydroxypentyloxy)cholic acid methyl ester; 3β-(hydroxyhexyloxy)cholic acid methyl ester; 3β-(hydroxydecanoyloxy)cholic acid methyl ester; 3β-(2-hydroxyethyloxyethyloxy)cholic acid methyl ester; 3β-(2-hydroxypropyloxy)cholic acid methyl ester; 3β-(hydroxyethyloxy)desoxycholic acid methyl ester; 3β-(hydroxypropyloxy)desoxycholic acid methyl ester; 3β-(hydroxypentyloxy)desoxycholic acid methyl ester; 3β-(hydroxydecyloxy)desoxycholic acid methyl ester; 3β-(2-hydroxyethyloxy)chenodesoxycholic acid methyl ester; 3β-(3-hydroxypropyloxy)chenodesoxycholic acid methyl ester; 3β-(5-hydroxypentyloxy)chenodesoxycholic acid methyl ester; 3β-(10-hydroxydecyloxy)chenodesoxycholic acid methyl ester; 3β-(2-hydroxyethyloxy)litocholic acid methyl ester; 3β-(3-hydroxypropyloxy)litocholic acid methyl ester; 3β-(5-hydroxypenthyloxy)lithocholic acid methyl ester; 3β-(10-hydroxydecayloxy)lithocholic acid methyl ester; 3β-(benzyloxyethyloxy)cholic acid methyl ester; 3β-(benzyloxyethyloxy)cholic acid tert.butyl ester; 3β-(2-hydroxyethyloxy)cholic acid tert.butyl ester; 3β-(2-hydroxyethyloxy)-7α,12α-diacetyloxycholic acid methyl ester; and 3β-(propionyloxy)-7α,12α-diacetyloxy-24-carboxylic acid methyl ester are mentioned as intermediates.

In the Swedish patent application with publication number 385,905, chenodeoxycholic acid is mentioned to have utility for the treatment of cholelithiasis and cholic acid is mentioned as an intermediate for the preparation thereof.

Swedish patent application with publication number 402,462 mentions sitosterol which may have medical application, e.g. for the prevention or reduction of absorption of cholesterol in the small intestine, and campesterol is mentioned to lower the effect of sitosterol.

In the Swedish patent application with publication number 413,247, 3α-hydroxycholestane and 3β-hydroxycholestane are mentioned to have antiinflammatoric properties and slightly side effects.

Swedish patent application with publication number 430,508 deals with compounds possibly having pharmacological properties, i.e. inhibition of HMG-CoA reductase and inhibi19tion of the formation of serum cholesterol. Examples of compounds specifically mentioned therein are 25-fluorocholest-5-en-3β,22-diol; 25-chlorocholest-5-en-3β,22-diol; 22-hydroxy-25-fluorocholest-5-en-3β-hemisuccinate; 22-hydroxy-25-chlorocholest-5-en-3β-hemisuccinate; and cholesta-5-en-3β,22,25-triol.

Compounds being known to stimulate the meiosis and being different from the compounds claimed in the present patent application are described in International patent applications having Nos. WO 96/00235, 96127658 and 97/00884 (Novo Nordisk A/S) and 98/55498. In International patent application having No. WO 98/52965, filed on May 11, 1998 and published on Nov. 16, 1998, it is stated that certain 20-aralkyl-5α-pregnan derivatives can be used in the preparation of a medicament for the control of fertility and some of the specific compounds mentioned therein are (3β,5α, 20R)-4,4,20-trimethyl-21-phenylpregna-8,14-dien-3-ol (example 1); (3β,5α,20R)-4,4,20-trimethyl-21-(3-methylphenyl)pregna-8,14-dien-3-ol (example 2A); and (3β,5α,20R)-4,4-dimethyl-23-phenyl-24-norchola-8,14-dien-3-ol (example 7A).

The compounds described herein have advantages compared with the known compounds.

SUMMARY OF THIS INVENTION

A main purpose of this invention is to furnish compounds which can be used to regulate meiosis.

One purpose of the present invention is to provide compounds and methods useful for relieving infertility in females and males, particularly in mammals, more particularly in humans.

In a further object, the present invention concerns the use of the compounds of the general formula Ic (stated in the claims, below) for relieving infertility in females and males, particularly in mammals, more particularly in humans.

In a further object of the present invention the compounds of the general formula I are useful as contraceptives in females and males, particularly in mammals, more particularly in humans.

In another embodiment, the invention relates to esters, salts, active metabolites and prodrugs of compound of the general formula Ia.

In still another preferred embodiment, the present invention relates to compounds of the general formula Ib (stated in the claims below) or esters, salts, active metabolites or prodrugs thereof as a medicament.

In a further preferred embodiment, this invention relates to compounds of the general formula Ic (stated in the claims below) or esters, salts, active metabolites or prodrugs thereof in the manufacture of a medicament for use in the regulation of meiosis.

In a further preferred aspect, the present invention relates to the use of a compound of formula Ib/Ic or an ester, salt, active metabolite or prodrug thereof as a medicament, in particular as a medicament for use in the regulation of meiosis. The compound may be used neat or in the form of a liquid or solid composition containing auxiliary ingredients conventionally used in the art.

In the present context, the expression “regulating the meiosis” is used to indicate that certain of the compounds of formula Ia, Ib or Ic can be used for stimulating the meiosis in vitro, in vivo, or ex vivo. Thus, the compounds of formula Ia, Ib or Ic which may be agonists of a naturally occurring meiosis activating substance, can be used in the treatment of infertility which is due to insufficient stimulation of meiosis in females and males. Other compounds of formula Ia, Ib or Ic, which may be antagonists of a naturally occurring meiosis activating substance, can be used for regulating the meiosis, preferably in vivo, in a way which makes them suited as contraceptives. In this case the “regulation” means partial or total inhibition.

In a still further preferred aspect, the present invention relates to the use of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof in the regulation of the meiosis of an oocyte, in particular a mammalian oocyte, more particularly a human oocyte.

In a still further preferred aspect, the present invention relates to the use of a compound of formula Ic organ ester, salt, active metabolite or prodrug thereof in the stimulation of the meiosis of an oocyte, in particular a mammalian oocyte, more particularly a human oocyte.

In a still further preferred aspect, the present invention relates to the use of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof in the inhibition of the meiosis of an oocyte, in particular a mammalian oocyte, more particularly a human oocyte.

In a still further preferred aspect, the present invention relates to the use of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof in the regulation of the meiosis of a male germ cell, in particular a mammalian male germ cell, more particularly a human male germ cell.

In a still further preferred aspect, the present invention relates to the use of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof in the stimulation of the meiosis of a male germ cell, in particular a mammalian male germ cell, more particularly a human male germ cell.

In a still further preferred aspect, the present invention relates to the use of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof in the inhibition of the meiosis of a male germ cell, in particular a mammalian male germ cell, more particularly a human male germ cell.

In a yet still further preferred aspect, the present invention relates to a method of regulating the meiosis in a mammalian germ cell which method comprises administering an effective amount of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof to a germ cell in need of such a treatment.

In a still further aspect, the present invention relates to a method of regulating the meiosis in a mammalian germ cell wherein a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof is administered to the germ cell by administering the compound to a mammal hosting said cell.

In a still further aspect, the present invention relates to a method wherein the germ cell the meiosis of which is to be regulated by means of a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof is an oocyte.

In a still further aspect, the present invention relates to a method of regulating the meiosis in an oocyte wherein a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof is administered to the oocyte ex vivo.

In a still further aspect, the present invention relates to a method of regulating the meiosis of a male germ cell by administering a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof to the cell.

In a still further aspect, the present invention relates to a method whereby mature male germ cells are produced by administering in vivo or in vitro a compound of formula Ic or an ester, salt, active metabolite or prodrug thereof to testicular tissue containing immature cells.

In a still further aspect, the present invention relates to compounds of formula Ia, Ib and Ic having improved stability.

According to the present invention there are provided novel compounds of formula Ia (stated in claim 1, below) with interesting pharmacological properties. The compounds described herein are useful for regulating the meiosis in oocytes and in male germ cells.

DETAILED DESCRIPTION OF THIS INVENTION

It has, surprisingly, been found that compounds having a side chain (R²²) which is different from the cholesterol and lanosterol side chains or compounds having certain specifically elected substituents in the ring system, have superior properties.

Preferred compounds of formula Ia, Ib and Ic are such having a double bond.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹ is methyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹ is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R², together with

R³, designates an additional bond between the carbon atoms at which R² and R³ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R² is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R² is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R² is C₁-C₈ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R² is C₁-C₃ alkoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R² is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is alkoxy, aralkyloxy, alkoxyalkoxy or alkanoyloxyalkyl, each group comprising a total of up to 10 carbon atoms, preferably up to 8 carbon atoms.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is C₁-C₄ alkoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is methoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is ethoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is CH₃OCH₂O—.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is pivaloyloxymethoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is an acyloxy group derived from an acid having from 1 to 20 carbon atoms.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is an acyloxy group selected from the group comprising acetoxy, benzoyloxy, pivaloyloxy, butyryloxy, nicotinoyloxy, isonicotinoyloxy, hemi succinoyloxy, hemi glutaroyloxy, butylcarbamoyloxy, phenylcarbamoyloxy, butoxycarbonyloxy, tert-butoxycarbonyloxy and ethoxycarbonyloxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is sulphonyloxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is phosphonyloxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ together with R′³ is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is the group ═NOH.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is a group of the general formula ═NOR³⁸, wherein R³⁸ is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is hydroxy and C₁-C₄ alkyl bound to the same carbon atom of the sterol skeleton.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R³ is perfluoro(lower alkyl), preferably perfluoro(lower alkyl) having 1 through 6, preferably 1 through 3, carbon atoms in the alkyl group, more preferred trifluoromethyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ and R′⁴ are both hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein one of R⁴ and R′⁴ is hydrogen while the other is methyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ and R′⁴ are both methyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ is branched or unbranched C₁-C₆ alkyl, optionally substituted by halogen, hydroxy or cyano.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′⁴ is branched or unbranched C₁-C₆ alkyl, optionally substituted by halogen, hydroxy or cyano.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ is hydroxy and R⁴ is selected from the group comprising hydrogen and branched or unbranched C₁-C₆ alkyl which may be substituted by halogen, hydroxy or cyano.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′⁴ is hydroxy and R⁴ is selected from the group comprising hydrogen and branched or unbranched C₁-C₆ alkyl which may be substituted by halogen, hydroxy or cyano.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ and R′⁴ together designate methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ and R′⁴ together with the carbon atom to which they are bound, form a cyclopropane ring.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ and R′⁴ together with the carbon atom to which they are bound, form a cyclopentane ring.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁴ and R′⁴ together with the carbon atom to which they are bound, form a cyclohexane ring.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is alkyl with 1 through 3 carbon atoms, preferably methyl, cyano or hydroxymethyl, or R⁵ is, together with R⁴, a methano bridge or R⁵ is, together with R⁴, an additional bond.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is a primary or secondary amide derived from a carboxylic acid.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is an ester with a C₁-C₆-alcohol group.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁶ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁶ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁶ is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁶ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁶, together with R⁵, designates an additional bond between the carbon atoms at which R⁵ and R⁶ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is methoxy or acetoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is the group ═NOH.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ is a group of the general formula ═NOR³⁶, wherein R³⁶ is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷ simultaneously is hydroxy and C₁-C₄ alkyl both being bound to the same carbon atom of the sterol skeleton, i.e. in the 7 position.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷, together with R⁹, designates an additional bond between the carbon atoms at which R⁷ and R⁶ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁷, together with R⁸, designates an additional bond between the carbon atoms at which R⁷ and R⁸ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁸, together with R⁹, designates an additional bond between the carbon atoms at which R⁸ and R⁹ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁸ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁸ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁸ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁹ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁹ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁹ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is methoxy or acetoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is the group ═NOH.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ is a group of the general formula ═NOR³⁷, wherein R³⁷ is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹ simultaneously is hydroxy and C₁-C₄ alkyl both being bound to the same carbon atom of the sterno skeleton, i.e. in the 11 position.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹, together with R⁹, designates an additional bond between the carbon atoms at which R¹¹ and R⁹ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹¹, together with R¹², designates an additional bond between the carbon atoms at which R¹¹ and R¹² are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is C₁-C₄ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is methoxy or acetoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is the group ═NOH.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹² is a group of the general formula ═NOR³³, wherein R³³ is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁴ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁴ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁴, together with R⁸, designates an additional bond between the carbon atoms at which R¹⁴ and R⁸ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is C₁-C₄ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is methoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is oxa.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is the group ═NOH.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵ is a group of the general formula ═NOR³², wherein R³² is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁵, together with R¹⁴, designates an additional bond between the carbon atoms at which R¹⁵ and R¹⁴ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is methoxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is the group ═NOH.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ is a group of the general formula ═NOR³⁴, wherein R³⁴ is C₁-C₃ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁶ together with R¹⁷, designates an additional bond between the carbon atoms at which R¹⁶ and R¹⁷ are placed.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁷ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁷ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹⁷ is in the α position.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²⁰ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²⁰ is hydroxymethyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²⁰ is C₁-C₄ alkyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²⁰ together with R′²⁰ designates methylene.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²⁰ together with R′²⁰ designates oxo.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′²⁰ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′²⁰ is halogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′²⁰ is methyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′²⁰ is hydroxy.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²² is cyclohexyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen, amino, cyano, carboxy, a group of the general formula —COOR³⁹, oxo, N-alkylamino or N,N-dialkylamino wherein the N-alkylamino or N,N-dialkylamino substituent optionally is substituted by carboxy, lower alkoxy or lower alkylthio; cyclohexylalkyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, lower alkoxy, halogen, amino, cyano, carboxy, a group of the general formula —COOR³⁹, oxo, N-alkylamino or N,N-dialkylamino wherein the N-alkylamino or N,N-dialkylamino substituent optionally is substituted by carboxy, lower alkoxy or lower alkylthio;

-   -   alkyl optionally substituted by one or more of the following         groups which substituents may be different or identical:         hydroxy, alkoxy, halogen, amino, cyano, carboxy, a group of the         general formula —COOR³⁹, oxo, N-acylamino, N-alkylamino or         N,N-dialkylamino wherein the N-alkylamino or N,N-dialkylamino         substituent optionally is substituted by carboxy, lower alkoxy         or lower alkylthio; or     -   alkenyl optionally substituted by one or more of the following         groups which substituents may be different or identical:         hydroxy, lower alkoxy, halogen, amino, cyano, carboxy, a group         of the general formula —COOR³⁹, oxo, N-alkylamino or         N,N-dialkylamino wherein the N-alkylamino or N,N-dialkylamino         substituent optionally is substituted by carboxy, lower alkoxy         or lower alkylthio;     -   and R³⁹ represents lower alkyl or aralkyl, e.g. benzyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R²² is phenyl, cyclohexyl, benzyl, o-tolyl, m-tolyl, p-tolyl, but-3-enyl, 3-methylbut-3-enyl, 2-methylpropyl, 2-oxo-2-ethoxyethyl, 2-oxo-2-(N,N-dimethylamino)ethyl, carboxymethyl, 3-hydroxymethylbutyl, 2-cyanoethyl, cyclohexyl methyl, 3-chloro-3-methylbutyl, 2-(N,N-dimethylamino)-2-cyanoethyl, 2-chloroethyl, 2-iodoethyl, ethyl, 2-phenylethyl, 2-methoxyethyl, 2-benzyloxyethyl or 2-acetoxyethyl.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R′²² is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein the side chain in the 17 position (i.e. —C(R²⁰) (R′²⁰)—CH(R²²)(R′²²)) is in the β position.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R² together with R³ is an additional double bond and R′³ is hydrogen.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹ is hydrogen, halogen, methyl, hydroxy or oxo; R² is selected from the group comprising hydrogen, hydroxy, C₁-C₃ alkyl, vinyl, C₁-C₃ alkoxy and halogen, or R² designates, together with R³, an additional bond between the carbon atoms at which R² and R³ are placed; R³ is selected from the group comprising hydrogen, hydroxy, optionally substituted alkoxy, acyloxy, sulphonyloxy, phosphonyloxy, oxo, halogen, C₁-C₄ alkyl and a group of the general formula ═NOR³⁸ wherein R³⁸ is hydrogen or C₁-C₃ alkyl, or R³ designates, together with R², an additional bond between the carbon atoms at which R² and R³ are placed; wherein R⁴ and R^(4′), which are different or identical with the proviso that they are not both hydroxy, are selected from the group comprising hydrogen, halogen, hydroxy and branched or unbranched C₁-C₆ alkyl which may be substituted by halogen, hydroxy or cyano, or wherein R⁴ and R^(4′) together designate methylene or oxo or, together with the carbon atom to which they are bound, form a cyclopropane ring, a cyclopentane ring, or a cyclohexane ring; R⁵ is hydrogen, halogen or hydroxy, or R⁵ designates, together with R⁶, an additional bond between the carbon atoms at which R⁵ and R⁶ are placed; R⁶ is hydrogen, hydroxy, halogen or oxo, or R⁶ designates, together with R⁵ or R⁷, an additional bond between the carbon atoms at which R⁶ and R⁵ or R⁷ are placed; R⁷ is selected from the group comprising hydrogen, methylene, hydroxy, methoxy, acetoxy, oxo, halogen, C₁-C₄ alkyl and a group of the general formula ═NOR³⁶ wherein R³⁶ is hydrogen or C₁-C₃ alkyl, or R⁷ designates, together with R⁶ or R⁸, an additional bond between the carbon atoms at which R⁷ and R⁶ or R⁸ are placed; R⁸ is hydrogen, hydroxy or halogen, or R⁸ designates, together with R⁷, R⁹ or R¹⁴, an additional bond between the carbon atoms at which R⁸ and R⁷, R⁹ or R¹⁴ are placed; R⁹ is hydrogen, hydroxy or halogen, or R⁹ designates, together with R⁸ or R¹¹, an additional bond between the carbon atoms at which R⁹ and R⁸ or R¹¹ are placed; R¹¹ is selected from the group comprising hydrogen, methylene, hydroxy, methoxy, acetoxy, oxo, halogen, C₁-C₄ alkyl and a group of the general formula ═NOR³⁷ wherein R³⁷ is hydrogen or C₁-C₃ alkyl, or R¹¹ designates, together with R⁹ or R¹², an additional bond between the carbon atoms at which R¹¹ and R⁹ or R¹² are placed; R¹² is selected from the group comprising hydrogen, halogen, C₁-C₄ alkyl, methylene, hydroxy, methoxy, acetoxy, oxo and a group of the general formula ═NOR³³ wherein R³³ is hydrogen or C₁-C₃ alkyl, or R¹² designates, together with R¹¹, an additional bond between the carbon atoms at which R¹¹ and R¹² are placed; R¹⁴ is hydrogen or hydroxy, or R¹⁴ designates, together with R¹⁵, an additional bond between the carbon atoms at which R¹⁴ and R¹⁵ are placed; R⁵ is selected from the group comprising hydrogen, halogen, C₁-C₄ alkyl, methylene, hydroxy, methoxy, oxo and a group of the general formula ═NOR³² wherein R³² is hydrogen or C₁-C₃ alkyl, or R¹⁵ designates, together with R¹⁴ an additional bond between the carbon atoms at which R¹⁵ and R¹⁴ are placed; R¹⁶ is selected from the group comprising hydrogen, halogen, C₁-C₃ alkyl, methylene, hydroxy, methoxy, oxo and a group of the general formula ═NOR³⁴ wherein R³⁴ is hydrogen or C₁-C₃ alkyl, or R¹⁶ designates, together with R¹⁷, an additional bond between the carbon atoms at Which R¹⁶ and R¹⁷ are placed; R¹⁷ is hydrogen or hydroxy, or R¹⁷ designates, together with R¹⁶, an additional bond between the carbon atoms at which R¹⁷ and R¹⁶ are placed; R²⁰ is selected from the group comprising hydrogen, C₁-C₄ alkyl and hydroxymethyl, or R²⁰ and R^(20′) together designate methylene or oxo; R^(20′) is hydrogen, halogen, alkyl or hydroxy, R^(220′) is hydrogen, hydroxy or oxo; R²² represents phenyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen (chloro, bromo or iodo), amino, N-alkylamino, N,N-dialkylamino, cyano, carboxy or oxo; benzyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen (chloro, bromo or iodo), amino, N-alkylamino, N,N-dialkylamino, cyano, carboxy or oxo; cyclohexyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen (chloro, bromo or iodo), amino, N-alkylamino, N,N-dialkylamino, cyano, carboxy or oxo; cyclohexylalkyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen (chloro, bromo or iodo), amino, N-alkylamino, N,N-dialkylamino, cyano, carboxy or oxo; alkyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen (chloro, bromo or iodo), amino, N-alkylamino, N,N-dialkylamino, cyano, carboxy or oxo; alkenyl optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxy, alkoxy, halogen (chloro, bromo or iodo), amino, N-alkylamino, N,N-dialkylamino, cyano, carboxy or oxo; and esters thereof.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R¹, R², R′⁴, R⁵, R¹¹, R¹², R¹⁵, R¹⁶, R′²⁰ and R′²² is each hydrogen, R³ is hydrogen, lower alkyl or perfluoro(lower alkyl), R′³ is hydroxy, or R³ designates, together with R′³, oxo, R⁴ is hydrogen or together with R⁵ a methano bridge or together with R⁵ an additional bond, R⁵ is lower alkyl, cyano, hydroxymethyl, a carbaldehyde, an oxime derived from a carbaldehyde, a carboxylic acid, a primary or secondary amide derived from a carboxylic acid, an ester with a C₁-C₆-alcohol group or together with R⁴ a methano bridge or together with R⁴ an additional bond, R⁷ is together with R⁸ an additional bond or hydrogen, if R⁸ and R⁹ or R⁸ and R¹⁴ stand together for an additional bond, R⁸ is together with R⁷ or with R⁹ or with R¹⁴ an additional bond, R⁹ is together with R⁸ an additional bond or a hydrogen atom, if R⁷ and R⁸ or R⁸ and R¹⁴ stand together for an additional bond, R¹⁴ is, together with R⁸, an additional bond or a hydrogen atom, if R⁷ and R⁸ or R⁸ and R⁹ stand together for an additional bond, R¹⁷ is hydrogen in the alpha position, R¹⁹ is methyl in the beta position, R²⁰ is methyl in the alpha position, and R²² is 3-methylbutyl; and ester, salt, active metabolite or prodrugs thereof.

Other preferred compounds of formula Ia, Ib and Ic are such wherein R⁵ is a C₁-C₃-alkyl group, preferably a methyl group, a cyano group, a hydroxymethyl group or together with R⁴ a methano bridge or together with R⁴ an additional bond.

It is to be understood that the above preferred substituents can be combined in any way with each other.

Examples of interesting and preferred compounds of the general formula Ia, Ib and Ic are as follows:

-   -   (20R)-20-Methyl-21-phenyl-5α-pregna-8,14-dien-3β-ol;         (20R)-20-methyl-21-(3-methylphenyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-20-methyl-21-(3-hydroxyphenyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-20-methyl-21-(cyclopentyl)-5α-pregna-8,14-dien-3β-ol;         24-nor-cholest-8,14-dien-3β-ol;         (20R)-20-methyl-21-(cyclohexyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-20-methyl-22-phenyl-5α-pregna-8,14-dien-3β-ol;         23,24-dinor-cholest-8,14-dien-3β-ol;         (20R)-20-methyl-21-(cyclobutyl)-5α-pregna-8,14-dien-3β-ol;         4,4-dimethyl-17β-((1R)-methyl-3-methyl-2-butenyl)         androsta-8,14-dien-3-β-ol;         (20R)-20-methyl-23-dimethylamino-5α-pregna-8,14-dien-3β-ol;         3β,hydroxy-5α-cycanochol-8-en-24-oic acid-N,N-dimethyl amide;         5β-methylchol-8-en-3-on -24-oic acid-N,N-dimethyl amide;         3β-hydroxy-4,4-dimethyl-5α,14β-chola-8,15-dien-24-oic         acid-N,N-dimethyl amide; 3β-hydroxy-5α-cyanocholest-8-en-24-one;         5β-methylchol-8-en-3,24-dione;         3β-hydroxy-4,4-dimethyl-5α,14β-cholesta-8,15-dien-24-one;         3β-hydroxy-5α-cycanochol-8-en-24-oic acid cyclohexyl ester;         5β-methylchol-8-en-3-on-24-oic acid cyclohexyl ester;         3β-hydroxy-4,4-dimethyl-5α,14β-chola-8,15-dien-24-oic acid         cyclohexyl ester; 3β-hydroxy-5α-chola-8,14-dien-24-oic         acid-N-(4-methylpiperazinyl)amide;         3β-hydroxychola-5,7-dien-24-oic         acid-N-(4-methylpiperazinyl)amide;         3β-hydroxy-5α-cyanochol-8-en-24-oic         acid-N-(4-methylpiperazinyl)amide;         5β-methylchol-8-en-3-on-24-oic         acid-N-(4-methylpiperazinyl)amide;         3β-hydroxy-4,4-dimethyl-5α,14β-chola-8,14-dien-24-oic         acid-N-(4-methylpiperazinyl)amide;         (20R)-20-methyl-21-phenyl-5α-pregna-5,7-dien-3β-ol;         (20R)-20-methyl-21-(3-methylphenyl)-5α-pregna-5,7-dien-3β-ol;         (20R)-20-methyl-21-(3-hydroxyphenyl)-5α-pregna-5,7-dien-3β-ol;         (20R)-20-methyl-21-(cyclopentyl)-5α-pregna-5,7-dien-3β-ol;         24-nor-cholest-5,7-dien-3β-ol;         (20R)-20-methyl-21-(cyclohexyl)-5α-pregna-5,7-dien-3β-ol;         (20R)-20-methyl-22-phenyl-5α-pregna-5,7-dien-3β-ol;         23,24-dinor-cholest-5,7-dien-3β-ol;         (20R)-20-methyl-21-(cyclobutyl)-5α-pregna-5,7-dien-3β-ol;         4,4-dimethyl-17β-((1R)-methyl-3-methyl-2-butenyl)androsta-5,7-dien-3β-ol;         (20R)-20-methyl-23-dimethylamino-5α-pregna-5,7-dien-3β-ol;         cholesta-5,7-dien-25-chloro-3β-ol;         cholesta-5,7-dien-26-chloro-3-β-ol; cholesta-5,7-dien-26-ol;         nor-24-cholesta-8,11-dien-3β-ol; cholesta-4,8-dien-3β-ol;         cholesta-4,8-dien-3α-ol; cholesta-4,8(14)-dien-3β-ol;         cholesta-4,8(14)-dien-3α-ol; 5-cyano-5α-cholest-7-en-3α-ol;         5-cyano-5α-cholest-7-en-3β-ol; 5-cyano-5β-cholest-7-en-3α-ol;         5-cyano-5α-cholest-8-en-3α-ol; 5-cyano-5α-cholest-8-en-3β-ol;         5-cyano-5β-cholest-8-en-3α-ol; 5-cyano-5β-cholest-8-en-3β-ol;         5-cyano-5α-cholest-8(14)-en-3α-ol;         5-cyano-5α-cholest-8(14)-en-3β-ol;         5-cyano-5β-cholest-8(14)-en-3α-ol;         5-cyano-5β-cholest-8(14)-en-3β-ol;         3′,4α-dihydrocyclopropa[4,5]-5β-cholest-7-en-3β-ol;         3′,4β-dihydrocyclopropa[4,5]-5α-cholest-7-en-3α-ol;         3′,4α-dihydrocyclopropa[4,5]-5β-cholest-8-en-3β-ol;         3′,4β-dihydrocyclopropa[4,5]-5α-cholest-8-en-3α-ol;         3′,4α-dihydrocyclopropa[4,5]-5β-cholest-8(14)-en-3β-ol;         3′,4β-dihydrocyclopropa[4,5]-5α-cholest-8(14)-en-3α-ol;         5-(hydroxymethyl)-5α-cholest-7-en-3β-ol;         5-(hydroxymethyl)-5β-cholest-7-en-3α-ol;         5-(hydroxymethyl)-5α-cholest-8-en-3β-ol;         5-(hydroxymethyl)-5β-cholest-8-en-3α-ol;         5-(hydroxymethyl)-5α-cholest-8(14)-en-3β-ol;         5-(hydroxymethyl)-5β-cholest-8(14)-en-3α-ol;         5-methyl-5β-cholest-8-en-3-one; 5-methyl-5β-cholest-8-en-3β-ol;         5-methyl-5β-cholest-8-en-3α-ol;         5-methyl-5β-cholest-8(14)-en-3-one;         5-methyl-5β-cholest-8(14)-en-3β-ol;         5-methyl-5β-cholest-8(14)-en-3α-ol;         3α-(trifluoromethyl)cholesta-4,7-dien-3β-ol;         3β-(trifluoromethyl)cholesta-4,7-dien-3α-ol;         3α-(trifluoromethyl)cholesta-4,8-dien-3β-ol;         3β-(trifluoromethyl)cholesta-4,8-dien-3α-ol;         3α-(trifluoromethyl)cholesta-4,8(14)-dien-3β-ol;         3β-(trifluoromethyl)cholesta-4,8(14)-dien-3α-ol;         5-methyl-24-nor-5β-cholest-8(14)-en-3-one;         (20R)-5,20-dimethyl-21-phenyl-5β-pregn-8(14)-en-3-one;         (20R)-21-cyclohexyl-5,20-dimethyl-5β-pregn-8(14)-en-3-one;         5-methyl-24-nor-5β-cholest-8(14),23-dien-3-one;         4,4-dimethyl-24-benzoylamido-5α-chola-8,14-dien-3β-ol;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid         N-phenylalanine amide;         mono(3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien)-24-succinate;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid         (1-methyl-4-hydroxypiperidinyl)ester;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(norleucine)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(arginine)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(glutamic acid)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(leucine) amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl         ester; 3β-hydroxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl         ester; 3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid         ethyl ester; 3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid; 3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid         cyclohexyl ester;         3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-24-one;         3β-hydroxy-4,4,24-trimethyl-5α-chola-8,14-dien-24-one;         3β-hydroxy-4,4-dimethyl-24-phenyl-5α-chola-8,14-dien-24-one;         3β-hydroxy-4,4-di-methyl-24-(3-pentyl)-5α-chola-8,14-dien-24-one;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-phenyl         amide; 3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid         amide; 4,4-dimethyl-24-phenylamino-5α-chola-8,14-dien-3β-ol;         4,4-dimethyl-24-amino-5α-chola-8,14-dien-3β-ol;         4,4-dimethyl-5α-chola-8,14-dien-3,24-diol;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-aldehyde;         4,4-dimethyl-17β-((1R)-methyl-4-methyl-3-pentenyl)androsta-8,14-dien-3β-ol;         4,4-dimethyl-5α-cholesta-14,16,24-triene-3β-ol;         4,4-dimethyl-17β-((1R)-methyl-3-methyl-2-butenyl)androsta-8,14-dien-3β-ol;         (20R)-4,4,20-trimethyl-21-(4-methylphenyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-4,4,20-trimethyl-21-(2-methylphenyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-4,4,20-trimethyl-21-(cyclohexyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-4,4,20-trimethyl-21-(3-hydroxyphenyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-4,4,20-trimethyl-22-(cyclohexyl)-5α-pregna-8,14-dien-3β-ol;         24-nor-4,4-dimethyl-5α-cholest-8,14-dien-3β-ol;         27-nor-4,4-dimethyl-5α-cholest-8,14,25-trien-3β-ol;         (20R)-4,4,20-trimethyl-21-(cyclobutyl)-5α-pregna-8,14-dien-3β-ol;         (20R)-4,4,20-trimethyl-21-(cyclopentyl)-5α-pregna-8,14-dien-3β-ol;         25-chloro-4,4-dimethyl-5α-cholesta-8,14-dien-3β-ol;         4,4-dimethyl-24-(N,N-dimethylamino)-24-cyano-5α-cholesta-8,14-dien-3β-ol;         4,4-dimethylcholest-8,14,25-trien-3β-ol;         4,4-dimethyl-17β-((1R)-methyl-4-iodobutyl)androsta-8,14-dien-3β-ol;         4,4-dimethyl-17β-((1R)-methylbutyl)androsta-8,14-dien-3β-ol;         4,4-dimethyl-17β-((1R)-methyl-4-cyanobutyl)androsta-8,14-dien-3β-ol;         4,4-dimethyl-17β-((1R)-methyl-4-cyanobutyl)androsta-8,14-dien-3β-ol;         27-nor-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26-oic acid         benzyl ester; 3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(methionine methyl ester)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(methionine)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(4-methylpiperazinyl) amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-tert-butylamide;         3,β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(isonipecotic acid ethyl ester)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(isonipecotic acid)amide;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic         acid-N-(phenylalanine methyl ester)amide;         3β-hydroxy-4,4-dimethylchola-5,7-dien-24-oic acid;         3β-hydroxy-4,4-dimethylchola-5,7-dien-24-oic acid-N-dimethyl         amide; 4,4-dimethyl-24-acetamido-5α-chola-8,14-dien-3β-ol;         4,4-dimethyl-24-acetoxy-5α-chola-8,14-dien-3β-ol;         4,4-dimethyl-24-methoxy-5α-chola-8,14-dien-3β-ol;         4,4-dimethyl-24-benzyloxy-5α-chola-8,14-dien-3β-ol;         3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid benzyl         ester;         26,27-diethyl-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioate;         3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioic acid;         and 27-nor-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26-oic         acid.

Preferred compounds of formula Ia, Ib and Ic are such which when tested by the method described below for agonistic properties (example 71) shows a relative activity of at least 50, preferably at least 80, or when tested by the method described below for antagonistic properties (example 72) shows a IC₅₀ value below 10, preferably below 2.

Examples of other preferred compounds are such not being active at the oestrogen receptor, and preferably compounds not being active at other hormone receptors.

Further preferred embodiments are mentioned in the appended claims.

As used in the present description and claims, a lower alkyl group—when used alone or in combinations—may be a straight or branched alkyl group. Preferably, said alkyl group contains not more than 6 carbon atoms. Examples of preferred alkyl groups are methyl, ethyl, propyl, isopropyl, butyl, tert-butyl, pentyl and hexyl, more preferred methyl, ethyl, propyl, isopropyl, butyl and tert-butyl, still more preferred methyl and ethyl. In a preferred embodiment of this invention, the alkyl group contains not more than 4 carbon atoms, preferably not more than 3 carbon atoms.

As used in the present description and claims, alkoxy designates a straight or branched alkoxy group, preferably containing not more than 6 carbon atoms, preferably not more than 4 carbon atoms, most preferred not more than 3 carbon atoms. Examples of preferred alkoxy groups are methoxy, ethoxy and propoxy, more preferred methoxy and ethoxy.

As used in the present description and claims, N-alkylamino is an alkyl group connected to an amino group. Preferably, said alkyl group is a lower alkyl group as defined above. Preferred N-alkylamino groups are methylamino and ethylamino.

As used in the present description and claims, N,N-alkylamino is two alkyl group which are the same or different and which are connected to an amino group. Preferably, said alkyl group are lower alkyl groups as defined above. Preferred N,N-alkylamino groups are N,N-dimethylamino, N,N-diethylamino and N-methyl-N-ethylamino.

As used in the present description and claims, the expression alkenyl designates a straight or branched alkenyl group preferably containing not more than 6 carbon atoms, preferably not more than 3 carbon atoms.

As used in the present description and claims, the expression halogen preferably designates chloro, bromo or iodide. Another group of preferred halogens are fluoro and chloro, preferably fluoro.

As used in the present description and claims, the expression C₃-C₆ cycloalkyl designates a cycloalkyl group containing 3 through 6 carbon atoms in the ring. Preferred examples are cyclopropyl and cyclopentyl.

As used in the present description and claims, the expression acyloxy designates a monovalent substituent comprising an optionally substituted C₁₋₄-alkyl or phenyl group linked through a carbonyloxy group; such as e.g. acetoxy, propionyloxy, butyryloxy, isobutyryloxy, pivaloyloxy, valeryloxy, benzoyl and the like. Optionally said acyloxy group may contain a hydroxy and/or carboxy group.

As used in the present description and claims, a statement that, e.g., R¹ is oxo means that oxo (═O) is present in the 1 position and, consequently, there is no hydrogen atom in the 1 position. Analogous considerations apply for similar situations. In other instances, two symbols together may represent oxo, e.g., R³ and R′³.

As used in the present description and claims, a statement that, e.g., R¹² is methylene means that methylene (═CH₂) is present in the 12 position and, consequently, there is no hydrogen atom in this position. Analogous considerations apply for similar situations. In other instances, two symbols together may represent methylene, e.g., R⁴ and R′⁴.

As used in the present description and claims, a statement that alkoxy is optionally substituted (R³) means that the alkoxy group is substituted with a convenient substituent such as hydroxy or carboxy.

As used in the present description and claims, the expressions “a primary or secondary amide derived from a carboxylic acid” used for the substituent R⁵ is a group of the general formula —CONHR⁴⁰ wherein R⁴⁰ is hydrogen or lower alkyl.

As used in the present description and claims, the expressions “an ester with a C₁-C₆-alcohol group” used for the substituent R⁵ is a group of the general formula —COOR⁴¹ wherein R⁴¹ is lower alkyl or aralkyl.

As used in the present description and claims, the expression aralkyl designates alkyl substituted by an aryl group, e.g. benzyl.

Salts of compounds of formula Ia, Ib and Ic are preferably pharmaceutically acceptable salts, especially acid-addition salts, including salts of organic acids and mineral acids. Examples of such salts include salts of organic acids such as formic acid, fumaric acid, acetic acid, propionic acid, glycolic acid, lactic acid, pyruvic acid, oxalic acid, succinic acid, malic acid, tartaric acid, citric acid, benzoic acid, salicylic acid and the like. Suitable inorganic acid-addition salts include salts of hydrochloric, hydrobromic, sulphuric and phosphoric acids and the like. Further examples of pharmaceutically acceptable inorganic or organic acid addition salts include the pharmaceutically acceptable salts listed in Journal of Pharmaceutical Science, 66 (1977), 2 et seq.

Esters of compounds of formula Ia, Ib and Ic are formally derived by esterification of one or more hydroxylic groups of a compound of formula Ia, Ib or Ic, respectively, with an acid which can for example be selected from the group of acids comprising succinic acid and other aliphatic dicarboxylic acids, nicotinic acid, isonicotinic acid, ethylcarbonic acid, phosphoric acid, sulphonic acid, sulphamic acid, benzoic acid, acetic acid, propionic acid and other aliphatic monocarboxylic acids.

A “metabolite” of a compound of formula Ia, Ib or Ic is an active derivative of a compound of formula Ia, Ib or Ic which is produced when the compound of formula Ia, Ib or Ic is metabolised. Metabolites of compounds of formula Ia, Ib or Ic can be identified either by administration of a compound of formula Ia, Ib or Ic to a host and an analysis of blood samples from the host, or by incubation of a compound of formula Ia, Ib or Ic with hepatic cells in vitro and analysis of the incubant.

A “prodrug” is a compound that either is converted into a compound of formula Ia, Ib or Ic in vivo or which has the same active metabolites as a compound of formula Ia, Ib or Ic.

The compounds of formula Ia, Ib or Ic have a number of chiral centres in the molecule and thus exists in several isomeric forms. All these isomeric forms and mixtures thereof are within the scope of the invention.

The compounds of the general formula Ia, Ib and Ic can be prepared analogously with the preparation of known compounds. Hence, synthesis of the compounds of formula Ia, Ib and Ic can followed the well established synthetic pathways described in the comprehensive sterol and steroid literature. The following books can be used as the key source in the synthesis: L. F. Fieser & M. Fieser: Steroids: Reinbold Publishing Corporation, NY 1959; Rood's Chemistry of Carbon Compounds (editor S. Coffrey): Elsevier Publishing Company, 1971; J. Fried and J. A. Edwards: Organic Reactions in Steroid Chemistry, Vol. I and II, Van Nostrand Reinhold Company, New York, 1972; and especially Dictionary of Steriods (editors: R. A. Hill; D. N. Kirk; H. L. J. Makin and G. M. Murphy): Chapmann & Hall: The last one contains an extensive list of citations to the original papers covering the period up to 1990. All these books including the last mentioned citations are incorporated by reference. In addition, information in all the above publications (including patent specifications) dealing with preparation of compounds similar with compounds of formula Ia, Ib and Ic is incorporated by reference.

Particularly, the compounds of the present invention may be synthesised according to the following general procedures:

Cholesta-5,8-dien-3-ol 1, which is synthesised as described in the literature [J. Lip. Res. 37, 1529, (1996)], can be oxidised in an Oppenauer reaction to give cholesta-4,8-dien-3-one 2 (scheme 1). In this reaction, the sterol is treated with a ketone like acetone, quinone or cyclohexanone in the presence of aluminum isopropoxide or aluminum tert-butoxide [e.g. J. Chem. Soc. Perkin 12667 (1994)]. The sterol can also be oxidised with pyridinium dichromate [vide Synth. Commun. 20 (1990), 1167]. The same oxidation reaction can be carried out with cholesta-5,8(14)-dien-3β-ol, which is also synthesised as described in the literature [J. Lip. Res. 37 (1996), 1529,] to give cholesta-5,8(14)-dien-3-one. For this ketone, a laborious synthesis is described in the literature [Bull. Soc. Chim. Fr. 2037, (1971)]. Cholesta-4,7-dien-3-one is synthesised according to literature procedures [Liebigs Ann. Chem. 542 (1939), 218,] (in scheme 1, the series with the Δ⁸-double bond are shown as an example, analogous reactions have to be performed in the Δ⁷- and Δ⁸⁽¹⁴⁾-series).

In the following, only the syntheses in the Δ⁸-series are described. The derivatives in the Δ⁷ and Δ⁸⁽¹⁴⁾-series can be synthesised by a skilled artisan from the corresponding starting materials in the same way.

Enone 2 can be treated with different C₁-C₆-Grignard reagents to give two diastereomeric alcohols 3 and 4 (with R³=C₁-C₆-alkyl), which are easily separated by column chromatography [e.g. J. Med. Chem. 40 (1997), 61].

Cholesta-4,8-dien-3-one 2 can be reduced according to well known literature procedures. Lithium aluminum hydride, sodium borohydride and diisobutylaluminum hydride are especially useful [e.g. Liebigs Ann. Chem. 542 (1939), 218]. Two diastereomeric alcohols of the formulae 3 and 4 (with R³=hydrogen) can be obtained and readily separated by column chromatography.

For the introduction of perfluoroalkyl substituents in position 3, cholesta-4,8-dien-3-one 2 can be treated with perfluoroalkyltrialkylsilanes in the presence of fluoride sources like tetrabutyl-ammonium fluoride or caesium fluoride. The trifluoromethyl group is preferentially introduced with reagents like trimethylsilyltrifluoromethane or triethylsilyltrifluoromethane [J. Org. Chem. 56 (1991), 984; J. Org. Chem. 54 (1989), 2873].

Again two diastereomeric alcohols of the formulae 3 and 4 (with R³ perfluoroalkyl, preferentially: trifluoromethyl) can be obtained and readily separated by column chromatography.

The cyanoketones 5 and 6 are available starting from cholesta-4,8-dien-3-one 2 via a conjugate addition of cyanide (scheme 2). Different reagents like diethylaluminum cyanide [J. Org. Chem. 59 (1994), 2766] and some alkali and earth alkali metal cyanides [Tetrahedron Lett. 28 (1987), 4189; Can. J. Chem. 59 (1981), 1641] can be used in this reaction.

The readily separated cyanoketones 5 and 6 can be reduced according to well known literature procedures. Lithium aluminum hydride, sodium borohydride and diisobutylaluminum hydride are used preferentially [e.g. Aust. J. Chem. 35 (1982), 629]. In the reduction reactions, two diastereomeric alcohols 7 and 8 (R³=H), respectively, 9 and 10 (R³=H) are obtained.

The cyanoketones of the formulae 5 and 6 can also be treated with Grignard reagents [e.g. Chem. Pharm. Bull. 9 (1961), 854] to give two diastereomeric tertiary alcohols 7 and 8 or 9 and 10 (with R³=C₁-C₆-alkyl), respectively.

The hydroxymethyl derivatives of the formulae 12 and 14 can be synthesised in two step sequences from the cyanoalcohols 8 and 9, respectively (scheme 3). First the cyano group can be reduced with an electrophilic reducing agent like diisobutylaluminum hydride to give the corresponding imines, which are hydrolysed in situ to the carbaldehydes 11 and 13. In the second step, thecarbaidehydes can be further reduced with well known reducing agents like lithium aluminum hydride, sodium borohydride or diisobutylaluminum hydride to the desired hydroxymethyl derivatives [e.g. J. Med. Chem. 39 (1996), 5092].

The methano derivatives of the formulae 15 and 16 can be synthesised form the allylic alcohols 3 and 4, respectively (scheme 4). Different variations of the Simmonsmith reaction can be employed. As reagents, diiodomethane in the presence of zinc/copper-couple [e.g. J. Org. Chem. 31 (1966), 3869; J. Med. Chem. 39 (1996), 4218] as well as chloroiodomethane in the presence of a diethylzinc solution [J. Am. Chem. Soc. 108 (1986), 6343] can be used in this cyclopropanation reaction.

5β-Methylcholest-8-en-3-one 17 can be synthesised from cholesta-4,8-dien-3-one 2 via a conjugate addition reaction (scheme 5). The methyl group is introduced either with lithium dimethylcuprate [e.g. Aust. J. Chem. 35 (1982), 629] or with methyl-Grignard compounds or trimethylaluminum in the presence of nickel catalysts [e.g. Tetrahedron Lett. 35 (1994), 6075; Synthesis 3 (1995), 317].

Subsequent reduction of 5β-methylketone 17 is achieved with different well known reducing agents like lithium aluminum hydride, sodium borohydride and diisobutylaluminum hydride [e.g. Aust. J. Chem. 35 (1982), 629]. Two diastereomeric alcohols 18 and 19 (R³=hydrogen) can be obtained and readily separated by column chromatography.

If 5β-methylketone 17 is treated with Grignard reagents, two diastereomeric tertiary alcohols (18 and 19; with R³=C₁-C₆-alkyl) are obtained, which are easily separated by column chromatography.

Analogues that combine a 5-cyano-substituent with different steroidal sidechains can be synthesized by the following general route (see scheme 6): Starting from lichesterol, which can be synthesized as described in the literature [J. Chem. Soc. Perkin Trans. 1 (1981), 2125], an enone can be generated by an oppenauer oxidation [e.g. J. Chem. Soc. Perkin Trans. 1 (1994), 2667]. A cyano group can be introduced by conjugate addition [J. Org. Chem. 59 (1994), 2766]. After protection of the 3-ketone as an ketal, the side chain can be cleaved by ozonolysis and subsequent reductive work up [Synthesis 3 (1990), 193]. The 22-alcohol can be transformed to the corresponding tosylate to generate the appropriate leaving group for the copper catalyzed addition of grignard reagents. By this addition, different alkyl- and aryl side chains can be introduced [Chem. Pharm. Bull. 28 (1980), 606]. After deprotection of the ketone, the latter can be reduced to the corresponding α- and β-alcohols by standard methods.

The compounds of the present invention will influence the meiosis in oocytes as well as in male germ cells.

The existence of a meiosis inducing substance in nature has been known for some time. However, until recently the identity of the meiosis inducing substance or substances was unknown.

The prospects of being able to influence the meiosis are several. According to a preferred embodiment of the present invention, a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof can be used to stimulate the meiosis. According to another preferred embodiment of the present invention, a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof can be used to stimulate the meiosis in humans. Thus, the compounds of formula Ia, Ib or Ic and ester, salt, active metabolite or prodrugs thereof are promising as new fertility regulating agents without the usual side effect on the somatic cells which are known from the hitherto used hormonal contraceptives which are based on estrogens and/or gestagens.

For use as a contraceptive agent in females, a meiosis inducing substance can be administered so as to prematurely induce resumption of meiosis in oocytes while they are still in the growing follicle, before the ovulatory peak of gonadotropins occurs. In women, the resumption of the meiosis can, for example, be induced a week after the preceding menstruation has ceased. When ovulated, the resulting overmature oocytes are then most likely not to be fertilised. The normal menstrual cycle is not likely to be affected. In this connection it is important to notice, that the biosynthesis of progesterone in cultured human granulosa cells (somatic cells of the follicle) is not affected by the presence of a meiosis inducing substance whereas the estrogens and gestagens used in the hitherto used hormonal contraceptives do have an adverse effect on the biosynthesis of progesterone.

According to another aspect of this invention, a meiosis inducing substance of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof can be used in the treatment of certain cases of infertility in females, including women, by administration thereof to females who, due to an insufficient own production of meiosis activating substance, are unable to produce mature oocytes. Also, when in vitro fertilisation is performed, better results can be achieved, when a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof is added to the medium in which the oocytes are cultured.

When infertility in males, including men, is caused by an insufficient own production of the meiosis activating substance and thus a lack of mature sperm cells, administration of a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof may relieve the problem.

As an alternative to the method described above, contraception in females can also be achieved by administration of a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof which inhibits the meiosis, so that no mature oocytes are produced. Similarly, contraception in males can be achieved by administration of a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof which inhibits the meiosis, so that no mature sperm cells are produced.

The route of administration of compositions containing a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof may be any route which effectively transports the active compound to its site of action.

Thus, when the compounds of this invention are to be administered to a mammal, they are conveniently provided in the form of a pharmaceutical composition which comprises at least one compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof in connection with a pharmaceutically acceptable carrier. For oral use, such compositions are preferably in the form of capsules or tablets.

From the above it will be understood that administrative regimen called for will depend on the condition to be treated. Thus, when used in the treatment of infertility, the administration may have to take place once only, or for a limited period, e.g. until pregnancy is achieved. When used as a contraceptive, the compounds of formula Ia, Ib or Ic or ester, salt, active metabolite or prodrugs thereof will either have to be administered continuously or cyclically. When used as a contraceptive by females and not taken continuously, the timing of the administration relative to the ovulation will be important.

Pharmaceutical Compositions

Pharmaceutical compositions comprising a compound of formula Ia, Ib or Ic or an ester, salt, active metabolite or prodrug thereof may further comprise carriers, diluents, absorption enhancers, preservatives, buffers, agents for adjusting the osmotic pressure, tablet disintegrating agents and other ingredients which are conventionally used in the art. Examples of solid carriers are magnesium carbonate, magnesium stearate, dextrin, lactose, sugar, talc, gelatin, pectin, tragacanth, methyl cellulose, sodium carboxymethyl cellulose, low melting waxes and cocoa butter.

Liquid compositions include sterile solutions, suspensions and emulsions. Such, liquid compositions may be suitable for injection or for use in connection with ex vivo and in vitro fertilisation. The liquid compositions may contain other ingredients which are conventionally used in the art, some of which are mentioned in the list above.

Further, a composition for transdermal administration of a compound of this invention may be provided in the form of a patch and a composition for nasal administration may be provided in the form of a nasal spray in liquid or powder form.

The dose of a compound of formula Ia, Ib or Ic to be used will be determined by a physician and will depend, inter alia, on the particular compound employed, on the route of administration and on the purpose of the use. In general, the compositions of the invention are prepared by intimately bringing into association the active compound with the liquid or solid auxiliary ingredients and then, if necessary, shaping the product into the desired formulation.

Usually, not more than 1000 mg, preferably not more than 100 mg, and in some preferred instances not more than 10 mg, of a compound of formula Ia, Ib or Ic is to be administered to mammals, e.g. to man, per day.

None of the compounds of formula Ia, Ib and Ic have shown to be toxic when administered to man in an amount of 1000 mg per day.

The compounds of formula Ia, Ib or Ic thereof can be synthesised by methods known per se.

The present invention is further illustrated by the following examples which, however, are not to be construed as limiting the scope of protection. The features disclosed in the foregoing description and in the following examples may, in any combination thereof, be material for realising the invention in diverse forms thereof.

EXAMPLE 1 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid Methyl Ester

4,4-Dimethyl-3-oxochol-5-en-24-oic acid methyl ester (19.8 g) (G. Aranda et al., Tetrahedron 43 (1987), 4147) is reduced with sodium borohydride (9.9 g) in 1900 ml methanol at room temperature. The solution is stirred for 20 hours. After aqueous work-up, 3β-hydroxy-4,4-dimethylchol-5-en-24-oic acid methyl ester (19.2 g) is isolated. Melting point: 136-138° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.55 (1H, m); 3.67 (3H, s); 3.23 (1H, m).

3β-Hydroxy-4,4-dimethylchol-5-en-24-oic acid methyl ester (19.1 g) is suspended in 390 ml of N,N-dimethylformamide (hereinafter designated DMF) and 39 g of imidazol and 34.6 g of tert-butyldimethylsilyl chloride is added. The reaction mixture is stirred at 70° C. for 20 hours. After aqueous work-up, a crude product 23.9 g is isolated. Crystallization from tetrahydrofurane/methanol (hereinafter tetrahydrofurane and methanol are designated THF and MeOH, respectively) yields 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-chol-5-en-24-oic acid methyl ester (19.55 g). Melting point: 144-146° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.52 (1H, m); 3.66 (3H, s); 3.18 (1H, m); 0.89 (9H, s); 0.02 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethylchol-5-en-24-oic acid methyl ester (9.75 g) is dissolved in a warm mixture of 122 ml of benzene and 475 ml of n-hexane, 1,3-dibromo-5,5-dimethylhydantoin (3.95 g) is added and the mixture is refluxed for 15 minutes. After cooling (fast) and evaporating to dryness under reduced pressure, 475 ml of o-xylene and 21.7 ml of quinaldine is added and the mixture is refluxed for 1 hour. After aqueous work-up and trituration with methanol 3β-tert-butyldimethylsilyloxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl ester (8.22 g) is isolated. Melting point: 116-124° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.88 (1H, m); 5.52 (1H, m); 3.65 (3H, s); 3.32 (1H, m); 0.89 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl ester (1.0 g) is suspended in a mixture of 25 ml methanol and 3.4 ml of concentrated hydrogen chloride acid and refluxed overnight. After evaporation to dryness under reduced pressure the remanens is triturated with methanol and thereafter recrystallized with methanol/water to give the title compound (0.47 g). Melting point: 156-157° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.36 (1H, s); 3.67 (3H, s); 3.23 (1H, m). MS (abbreviation for mass spectroscopy): Calculated: 414.6. Found 414.3.

EXAMPLE 2 3β-Hydroxy-4,4-dimethylchola-5,7-dien-24-oic Acid Methyl Ester

3β-tert-Butyldimethylsilyloxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl ester (0.50 g) is dissolved in 5 ml of dry THF and 0.48 g of tetra-butylammoniumfluoride hydrate and 0.4 g of pulverised molecular sieve is added and the mixture is stirred for 4 days. After column chromatography and crystallization from methanol/water and methanol, the title compound (195 mg) is isolated. Melting point: 124-128° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.91 (1H, m); 5.53 (1H, m); 3.67 (3H, s). 3.4 (1H, m). MS: Calculated: 414.6. Found 414.3.

EXAMPLE 3 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid Ethyl Ester

3β-tert-Butyldimethylsilyloxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl ester (4.0 g) is suspended in a mixture of 60 ml of 96% ethanol, 10.1 ml of benzene and 10.1 ml of concentrated hydrogen chloride acid and refluxed for 3.5 hours. After aqueous work-up and column chromatography followed by crystallization from ethanol the title compound (1.37 g) is isolated. Melting point: 122-124° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 4.13 (2H, q); 3.25 (1H, m). 1.25 (3H, t). MS: Calculated: 428.7. Found 428.3.

EXAMPLE 4 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid ethyl ester (1.9 g) is suspended in a mixture of 190 ml of 96% ethanol and 63 ml of 1M sodiumhydroxide. The mixture is stirred for 3.5 h at room temperature. After aqueous work-up and crystallization from ethanol/water, the title compound (1.48 g) is isolated. ¹H-NMR (CDCl₃, 400 MHz): δ=5.36 (1H, s); 3.22 (1H, m); 1.03 (3H, s). 1.01 (3H, s); 0.97 (3H, d); 0.83 (3H, s); 0.81 (3H, s). MS: Calculated: 400.6. Found 400.3.

EXAMPLE 5 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid Cyclohexyl Ester

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (0.2 g) is suspended in 10 ml of cyclohexanol, 0.1 ml of boron trifluoride diethyl etherate is added and the mixture is stirred at 55-60° C. for 20 hours. After evaporation to dryness under reduced pressure, the remanens is purified by column chromatography and crystallized from methanol to give the title compound (49 mg). Melting point: 130-132° C. ¹H-NMR (CDCl₃): δ=5.35 (1H, s); 4.75 (1H, m); 3.24 (1H, m). MS: Calculated: 482.8. Found 482.4.

EXAMPLE 6 3β-Hydroxy-4,4-dimethyl7-56-cholesta-8,14-dien-24-one

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl ester (18.2 g) is suspended in 400 ml of DMF and 14.8 g of imidazol and 13.0 g of t-butyldimethylsilylchloride is added. The reaction is stirred at 65° C. for 20 hours. After aqueous work-up and crystallization from ether/methanol, 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl/ester (21 g) is obtained. Melting point: 124-125° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.34 (1H, s); 3.68 (3H, s); 3.2 (1H, m); 0.9 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl ester (12 g) is dissolved in a mixture of 500 ml THF and 400 ml ethanol. 60 ml 1M sodium hydroxide is added and the mixture is stirred for 20 hours. After aqueous work-up and crystallization from ethanol/water, 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid. (11.26 g) is obtained. Melting point: 181-184° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.34 (1H, s); 3.19 (1H, m); 0.9 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (5.0 g) is dissolved in 250 ml of dry dichloromethane. After cooling to −15° C., 2.14 ml of N-methylmorpholine and 1.27 ml isobutylchloroformate is added and the mixture is stirred at −15° C. for 20 minutes, whereupon 0.98 g of N,O-dimethylhydroxy/amine, HCl is added and the mixture is stirred overnight and the temperature is slowly elevated to room temperature. After aqueous work-up, 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-methoxy-N-methyl amide (5.37 g) is obtained. Melting point: 134-135° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 3.69 (3H, s); 3.2 (1H, m); 3.18 (3H, s); 0.9 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-methoxy-N-methyl amide (0.57 g) is dissolved in 10 ml of dry THF and added to a solution of 10 mmol isopropylmagnesium bromide in 10 ml THF, whereupon it was cooled on an ice-bath. The mixture is stirred for some h and left overnight at 5° C. After aqueous work-up, column chromatography and crystallization from methanol, 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-cholesta-8,14-dien-24-one (238 mg) is obtained. Melting point: 120-122° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 3.2 (1H, m); 2.61 (1H, m); 0.9 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-cholesta-8,14-dien-24-one (0.1 g) is dissolved in 5 ml of ethanol, 0.2 ml of 6N hydrogen chloride is added and the mixture is stirred for 2 days. After aqueous work-up, column chromatography and crystallization from ethanol/water, the title compound (64 mg) is obtained. ¹H-NMR (CDCl₃, 400 MHz): δ=5.35 (1H, s); 3.24 (1H, m); 2.62 (1H, m). MS: Calculated: 426.7. Found 426.3.

EXAMPLE 7 3β-Hydroxy-4,4,24-trimethyl-5α-chola-8,14-dien-24-one

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24 oic acid-N-methoxy-N-methyl amide (0.57 g) is reacted with methylmagnesiumbromide and hydrolysed with ethanol/HCl following the procedure outlined in example 6 to give the title compound (0.20 g). Melting point: 183-185° C. ¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 3.24 (1H, m); 2.17 (3H, s). MS: Calculated: 398.6. Found 398.3.

EXAMPLE 8 3β-Hydroxy-4,4-dimethyl-24-phenyl-5α-chola-8,14-dien-24-one

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24 oic acid-N-methoxy-N-methyl amide (0.57 g) is reacted with phenylmagnesiumbromide and hydrolysed with ethanol/HCl following the procedure outlined in example 6 to give the title compound (0.31 g). Melting point: 196-199° C. ¹H-NMR (CDCl₃, 300 MHz): δ=8.0-7.4 (5H, m); 5.35 (1H, s); 3.24 (1H, m); 3.0 (2H, m). MS: Calculated: 460.7. Found 460.3.

EXAMPLE 9 3β-Hydroxy-4,4-dimethyl-24-(3-pentyl)-5α-chola-8,14-dien-24-one

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24 oic acid-N-methoxy-N-methyl amide (0.57 g) is reacted with 3-pentylmagnesiumbromide and hydrolysed with ethanol/HCl following the procedure outlined in example 6 to give the title compound (13 mg). ¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 3.23 (1H, m). MS: Calculated: 454.7. Found: 454.3.

EXAMPLE 10 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-phenyl Amide

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24 oic acid (0.50 g) is dissolved in 15 ml of dry dichloromethane. After cooling to −15° C., 0.188 ml of N-methylmorpholine and 0.153 ml of isobutylchloroformiate is added and the mixture is stirred at −15° C. for 20 minutes, whereupon 0.44 ml of aniline is added. The mixture is stirred overnight and the temperature is slowly elevated to room temperature. After aqueous work-up and crystallization from methanol, 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-phenyl amide (0.431 g) is obtained. H-NMR (CDCl₃, 400 MHz): δ=7.53 (2H, d); 7.34 (2H, t); 7.17 (1H, s); 7.12 (1H, t); 5.35 (1H, s); 3.21 (1H, m); 0.9 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-phenyl amide (50 mg) is dissolved in 5 ml of ethanol, 0.2 ml of 6N hydrogen chloride is added and the mixture is stirred at room temperature overnight. After aqueous work-up and crystallization from methanol the title compound is obtained. (36 mg). H-NMR (CDCl₃, 400 MHz): δ=7.53 (2H, d); 7.33 (2H, t); 7.18 (1H, s); 7.12 (1H, t); 5.36 (1H, s); 3.26 (1H, m). MS: Calculated: 475.7. Found: 475.4.

EXAMPLE 11 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid Amide

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24 oic acid (1.0 g) is reacted with ammonia and hydrolysed with ethanol/HCl following the procedure outlined in example to give the title compound (147 mg). Melting point: 233-235° C. ¹H-NMR (CDCl₃, 400 MHz)): δ=5.36 (1H, s); 5.45-5.2 (2H, broad d); 3.25 (1H, m). MS: Calculated: 399.6. Found 399.3.

EXAMPLE 12 4,4-Dimethyl-24-phenylamino-5α-chola-8,14-dien-3β-ol

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24 oic acid (0.15 g) is reacted with aniline following the procedure outlined in example 10 and reduced with lithium aluminum hydride (0.15 g) in THF at room temperature. Aqueous work-up and crystallization from methanol gives 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-24-phenylamino-5α-chola-8,14-dien (106 mg). ¹H-NMR (CDCl₃, 300 MHz): δ=7.17 (2H, t); 6.69 (1H, t); 6.6 (2H, d); 5.35 (1H, s); 3.6 (1H, s); 3.2 (1H, m); 3.09 (2H, m); 0.9 (9H, s); 0.04 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-24-phenylamino-5α-chola-8,14-dien (100 mg) is hydrolysed with ethanol/HCl at 50° C. Aqueous work-up and crystallization from ethanol gives the title compound (53 mg). Melting point: 178-180° C. ¹H-NMR (CDCl₃, 300 MHz)): δ=7.19 (2H, t); 6.7 (1H, t); 6.63 (2H, d); 5.36 (1H, s); 3.6 (1H, s); 3.26 (1H, m); 3.1 (2H, m). MS: Calculated: 461.7. Found: 461.3.

EXAMPLE 13 4,4-Dimethyl-24-amino-5α-chola-8,14-dien-3β-ol

The compound is synthesised following the procedure outlined in example 12. ¹H-NMR (DMSO-d₆, D₂O+HCl): δ=5.3 (1H, s); 3.03 (1H, m); 2.77 (2H, m). MS: Calculated: 385.6. Found: 385.3.

EXAMPLE 14 4,4-Dimethyl-5α-chola-8,14-dien-3β,24-diol

4,4-Dimethyl-3-oxochol-5-en-24-oic acid methyl ester (6.0 g) (G. Aranda et al., Tetrahedron 43 (1987), 4147) is reduced with lithium aluminum hydride (3.3 g) in THF (600 ml). After aqueous work-up and crystallization from diethyl ether, 4,4-dimethyl-5-ene-3β,24-diol (5.21 g) is obtained. Melting point: 201-202° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.55 (1H, m); 3.61 (2H, m); 3.23 (1H, m). MS: Calculated: 388.6. Found: 388.4.

A mixture of 4,4-dimethylchol-5-ene-3β,24-diol (57 g), imidazole (125 g) and tert-butyldimethylsilyl chloride (110.5 g) in DMF is stirred at 70° C. for 20 hours. After aqueous work-up and crystallization from methanol, 3β,24-bis(tert-butyldimethylsilyloxy)-4,4-dimethylchol-5-ene (87.7 g) is obtained. Melting point: 161-162° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.53 (1H, m); 3.58 (2H, m); 3.21 (1H, m); 0.9 (18H, m); 0.03 (12H, m).

3β,24-Bis(tert-butyldimethylsilyloxy)-4,4-dimethylchol-5-ene (44 g) is dissolved in a warm mixture of hexane (2 l) and benzene (540 ml). 1,3-Dibromo-5,5-dimethylhydantoin (15.32 g) is added and the mixture is refluxed for 20 min and then cooled rapidly to room temperature, and the insoluble material is removed by filtration. The filtrate is concentrated under reduced pressure and o-xylene (2 l) and quinaldine (84 ml) is added. The mixture is refluxed for 1 hour. After aqueous work-up and trituration with methanol, 3β,24-bis (tert-butyldimethylsilyloxy)-4,4-dimethylchola-5,7-diene (39 g) is isolated. Melting point: 98-106° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.9 (1H, m); 5.54 (1H, m); 3.58 (2H, m); 3.35 (1H, m); 0.91 (18H, s); 0.05 (12H, m).

A mixture of 3β,24-bis(tert-butyldimethylsilyloxy)-4,4-dimethylchola-5,7-diene (18.8 g) in 99.9% ethanol (375 ml), benzene (55 ml) and concentrated hydrochloric acid (55 ml) is heated to reflux for 4 hours. After standing overnight at room temperature, the reaction mixture is concentrated under reduced pressure. Crystallization of the remanens from ethanol/water gives 8.44 g of the title compound. Melting point: 203-208° C. ¹H-NMR (CDCl₃, 400 MHz): δ=5.36 (1H, s); 3.62 (2H, m); 3.23 (1H, m).

EXAMPLE 15 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-aldehyde

4,4-Dimethyl-5α-chola-8,14-dien-3β,24-diol (0.21 g) is dissolved in benzene (21 ml). Tris(triphenylphosphine) ruthenium(II)chloride (1.04 g) is added and the mixture is stirred at room temperature for 20 hours. Purification by column chromatography gives the title compound (129 mg). ¹H-NMR (CDCl₃, 300 MHz): δ=9.79 (1H, s); 5.36 (1H, s); 1.05 (3H, s); 1.01 (3H, s); 0.95 (3H, d); 0.82 (3H, s); 0.80 (3H, s).

EXAMPLE 16 4,4-Dimethyl-17β-((1R)-methyl-4-methyl-3-pentenyl)androsta-8,14-dien-3β-ol

¹H-NMR (CDCl₃, 400 MHz): δ=5.35 (1H, s); 5.18 (1H, t); 3.24 (1H, dd), 1.7 (3H, s); 1.6 (3H, s).

EXAMPLE 17 4,4-Dimethyl-5α-cholesta-14,16,24-triene-3β-ol

¹H-NMR (CDCl₃, 400 MHz): δ=5.98 (1H, m); 5.8 (1H, m); 5.12 (1H, t), 3.24 (1H, m); 1.7 (3H, s); 1.58 (3H, s).

EXAMPLE 18 4,4-Dimethyl-17β-((1R)-methyl-3-methyl-2-butenyl)androsta-8,14-dien-3β-ol

¹H-NMR (CDCl₃, 400 MHz): δ=5.35 (1H, s); 4.93 (1H, d); 3.24 (1H, m), 1.69 (3H, s); 1.65 (3H, s).

EXAMPLE 19 (20R)-4,4,20-Trimethyl-21-phenyl-5α-pregna-8,14-dien-3β-ol

19A: According to Steroids 26 (1975), p. 339-357, the 4,4-dimethylstigmasterole has been synthesized.

19B: Ozonolysis of compound 19A at −70° C. by slowly introduction of O₃, and following reduction at −70° C. by sodium-bis-(2-methoxyethoxy)aluminum hydride and further reduction at a temperature about −30° C. with lithium aluminium hydride gave afforded 23-nor-4,4-di-methyl-5α-cholest-5-en-3β-22-diol, which was diacetylated in pyridine with acetic acid anhydride. (1B).

19C: Compound 19B was isomerised by reflux in ethanol/6M hydrochloric acid giving 23-nor-4,4-dimethyl-5α-cholest-8,14-dien-3β,22-diol.

19D: Compound 19C was selectively C-22 tosylated by treatment of p-toluene sulfonyl chloride in pyridine by standing over night in room temperature. The compound was purified by colom chromatography and crystalised.

NMR: H ppm: 0.78 s CH₃; 0.82 s CH₃; 0.98 d CH₃; 1.02 s CH₃; 2.46 s CH₃-aromat; 3.22 m H 3 a: 5.38 s H15; 7.33 d 2H; 7,78 d 2H.

This intermediate, 19D, is used in several of the following examples.

19E: Compound 19D was reacted with phenylmagnesium bromide catalysed by Li₂CuCl₄ giving the title compound. (Chem. Pharm. Bull. 28 (1980), p. 606-611; Masuo Monsaki et al.).

NMR: H ppm: 0.82 d CH₃; 0.83 s CH₃; 1.02 s CH₃; 1.03 s CH₃; 2.55 m 1H; 2,91 dd 1H; 3.25 m 3 a H; 5.4a 1H (15); 7.17 m 2H; 7.28 m 3H.

EXAMPLE 20 (20R)-4,4,20-Trimethyl-21-(3-methylphenyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with 3-methylphenyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H ppm: 0.80 d CH₃; 0.82 2s 2CH₃; 1.0 2s 2CH₃; 2.33 s CH₃-aromat; 3.27 m H3 a 5.40 s 1H; 6.98 m 3H; 7.17 1H.

EXAMPLE 21 (20R)-4,4,20-Trimethyl-21-(4-methylphenyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with 4-methylphenyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H ppm: 0.84 s, 2d 3 CH₃; 1.04 2s 2CH₃; 2.3.3 s CH₃-aromat; 3.25 m H3 a; 5.40 s 1H; 7.05 m 4H.

EXAMPLE 22 (20R)-4,4,20-Trimethyl-21-(2-methylphenyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with 2-methylphenyl magnesium bromide and Li₂CuCl₄ giving the title compound. NMR: H ppm: 0.84 s+2d, 3 CH₃; 1.04 2s 2CH₃; 2.32 s CH₃-aromat; 3.25 m H3 a; 5.40 s 1H; 7.10 m 4H;

EXAMPLE 23 (20R)-4,4,20-Trimethyl-21-(cyclohexyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with cyclohexyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H: mmp: 0.81 s CH₃; 0.82s CH₃; 0.90 d CH₃; 1.03s CH₃; 104s CH₃; 3.24 s H3 a; 5.37 s H; MS: 424.4.

EXAMPLE 24 (20R)-4,4,20-Trimethyl-22-(phenyl)-5α-pregna-8,14-dien-3,3β-ol

According to example 19 (compound 19D) was reacted with tolyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H; ppm: 0.80 s CH₃; 0.82 s CH₃; 1.02s CH₃; 1.04 s CH₃; 1.06 d CH₃; 3.25 m H3 a; 5.35s H; 7.18 m 3H; 7.27 m 2H.

EXAMPLE 25 (20R)-4,4,20-Trimethyl-21-(3-hydroxyphenyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with 3-trimethylsilyloxyphenyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H; ppm: 0.80 d CH₃; 0.83 s CH₃; 0.84 s CH₃; 1.01 s CH₃; 1.03 s CH₃; 3.26 m H3 a; 5.40 s H; 6.66 m 2H; 6.76 m H; 7.16 m H. MS: 434.3.

EXAMPLE 26 (20R)-4,4,20-Trimethyl-22-(cyclohexyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with cyclohexylmethyl magnesium bromide and CuLi₂Cl₄ giving the title compound.

NMR: H ppm: 0.80 s CH₃; 0.83 s CH₃; 0.91 d CH₃; 1.01 s CH₃; 1.03 S CH₃; 3.24 m H3 a; 5.35 s H. MS: 483.3.

EXAMPLE 27 24-Nor-4,4-dimethyl-5α-cholest-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with isobutyl magnesium bromide and CuLi₂Cl₄ giving the title compound. NMR: H; ppm: 0.80 s CH₃; 0.84 s CH₃; 0.86 d CH₃; 0.88 d CH₃; 0.91 d CH₃; 1.02 s CH₃; 1.04 s CH₃; 3.25 m H3 a; 5.35 s H.

EXAMPLE 28 27-Nor-4,4-dimethyl-5α-cholesta-8,14,25-trien-3β-ol

According to example 19 (compound 19D) was reacted with cyclopropylmethyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H; ppm: 0.82 s CH₃; 0.84 S CH₃; 0.95 d CH₃; 1.02 s CH₃; 1.04 s CH₃; 3.24 H3 a; 4.97 dd 2H; 5.37 s 1H; 5.82 m 1H.

EXAMPLE 29 (20R)-4,4,20-Trimethyl-21-(cyclobutyl)-5α-pregna-8,14-dien-3β-ol

In the reaction of example 28 there was further isolated a compound which was identified as the title compound.

NMR: H; ppm: 0.80 s CH₃; 0.84 s CH₃; 0.89 d CH₃; 1.01 s CH₃; 1.03 s CH₃; 3.25 m H3 a; 5.35 s 1H.

EXAMPLE 30 (20R)-4,4,20-Trimethyl-21-(cyclopentyl)-5α-pregna-8,14-dien-3β-ol

According to example 19 (compound 19D) was reacted with cyclopentyl magnesium bromide and Li₂CuCl₄ giving the title compound.

NMR: H; ppm: 0.80 s CH₃; 0.82 5 CH₃; 0.94 d CH₃; 1.01 s CH₃; 1.03 s CH₃; 3.25 m H3 a; 5.35 s H.

EXAMPLE 31 25-Chloro-4,4-dimethyl-5α-pregna-8,14-dien-3β-ol

A mixture of 3β-(tert-butyldimethylsilyloxy)-4,4-dimethylcholesta-5,7,25-triene (50 mg), benzene (5 mL), ethanol (5 mL) and concentrated HCl (2 mL) was heated under reflux for 4 hours. The reaction was concentrated to half volume under reduced pressure and water (5 mL) was added. Extraction of the aqueous phase with dichloromethane and concentration under reduced pressure gave a residue, which was purified by flash chromatography. Recrystallisation from ethyl acetate:hexane gave the title compound as a white solid (35 mg). Melting point 131-133° C.

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.78 (s), 0.80 (s, 3H), 0.92 (d), 0.99 (s), 1.02 (s), 1.52 (s), 1.55 (s), 3.22 (dd), 5.33. The mass spectrum showed characteristic peaks at: 446:3 (M⁺).

EXAMPLE 32 4,4-Dimethyl-24-(N,N-dimethylamino)-24-cyano-5α-cholesta-8,14-dien-3β-ol

To a mixture of 3β-hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-aldehyde (10 mg, 0.03 mmol), dimethylamine hydrochloride (18 mg), sodium acetate (16 mg) and molecular seives (100 mg) in methanol (2 mL) was added sodium cyanoborohydride (8 mg) and the reaction stirred for 1.5 hours. Dilute HCl (0.2 mL) was added, followed by sodium bicarbonate solution (0.3 mL) and the seives were removed by filtration. Concentration of the remaining solution gave a residue which was purified by flash chromatography to give the title compound (5 mg).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.80 (s, 3H), 0.82 (s, 3H), 0.95 (d), 1.01 (s, 3H), 1.02 (s, 3H), 2.30 (s, 6H), 3.20 (dd, 1H), 3.39-3.49 (dd, 1H), 5.32 (s, 1H). The ¹³C-NMR spectrum (CDCl₃, d) showed characteristic signals at: 151.3, 142.3, 123.2, 117.5, 117.4, 79.1. The mass spectrum showed characteristic peaks at: 438.4 (M⁺).

EXAMPLE 33 4,4-Dimethylcholest-8,14,25-trien-3β-ol

Step 1

A solution of (25R)-4,4-dimethyl-5α-cholesta-8,14-diene-3β,26-diol (69 mg, 0.16 mmol), toluene sulphonyl chloride (45 mg, 2.4 mmol) and pyridine 1.5 ml was stirred for 6 h at ice bath temperature and 3 h at room temperature. Concentration under reduced pressure and purification by flash chromatography gave (25R)-(26-tosyloxy)-4,4-dimethyl-5α-cholesta-8,14-diene-3β-ol (61 mg).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.75 (s), 0.82 (s), 0.84 (s), 0.86 (s), 1.00 (s), 1.03 (s), 1.20 (s), 2.42 (s, 3H), 3.20-3.30 (m, 1H), 3.62-3.80 (m, 2H), 5.33 (s, 1H), 7.32 (d, 2H), 7.75 (d, 2H).

Step 2

(25R)-(26-Tosyloxy)-4,4-dimethyl-5α-cholesta-8,14-diene-3β-ol (61 mg), sodium iodide (150 mg and dimethylformamide (2 mL) was heated at 60° C. for 4 hours. Water was added and the aqueous layer extracted with dichloromethane, concentration under reduced pressure gave (25R)-26-iodo-4,4-dimethyl-5α-cholesta-8,14-dien-3β-ol, which was dissolved in chloroform (5 mL). 1,8-Diazabicyclo(5.4.0)undec-7-ene was added and the solution was heated to reflux for 5 h, concentrated under reduced pressure and purified by flash chromatography. Recrystallisation from methanol gave the title compound (5 mg).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.80 (s), 0.82 (s), 0.93 (d), 1.00 (s), 1.02 (s), 1.51 (s), 1.71 (s), 3.20-3.30 (m, 1H), 4.62 (d, 2H), 5.33 (s, 1H).

EXAMPLE 34 4,4-bimethyl-17β-((1R)-methyl-4-chlorobutyl) androsta-8,14-dien-3β-ol

Step 1

A mixture of 4,4-dimethyl-5α-chola-8,14-diene-3β-24-diol (900 mg, 23 mmol), tosyl chloride (470 mg, 24 mmol) in pyridine (20 mL) was stirred 6 h at ice bath temperature, poured into water and extracted with diethyl ether. Drying over magnesium sulphate, concentration under reduced pressure and purification by flash chromatography gave 24-toluene-sulphonyloxy-4,4-dimethyl-5α-chola-8,14-diene-3-ol (1.1 g).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.79 (s, 3H), 0.83 (s, 3H), 0.90 (d, 3H), 1.01 (s, 3H), 1.02 (s, 3H), 2.42 (s, 3H), 3.21 (dd, 1H), 4.01 (t, 2H), 5.33 (s, 1H), 7.35 (d, 2H), 7.80 (d, 2H).

Step 2

A mixture of 24-toluenesulphonyloxy-4,4-dimethyl-5α-chola-8,14-diene-3-ol (45 mg, 0.8 mmol) and lithium chloride (0.6 mmol) in dimethylformamide (1.5 mL) under nitrogen was heated at 60° C. for 2 hours. The reaction was then poured into water and extracted with diethyl ether. Drying over magnesium sulphate, concentration under reduced pressure and purification by flash chromatography followed by recrystallised from methanol gave the title compound, 12 mg.

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.80 (s, 3H), 0.82 (s, 3H), 0.97 (d, 3H), 1.02 (s, 3H), 1.04 (s, 3H), 3.25 (dd, 1H), 3.52 (m, 2H). The mass spectrum showed characteristic peaks at: 404.2 (M⁺).

EXAMPLE 35 4,4-Dimethyl-17β-((1R)-methyl-4-iodobutyl) androsta-8,14-dien-3β-ol

A mixture of 24-toluenesulphonyloxy-4,4-dimethyl-5α-chola-8,14-diene-3-ol (100 mg, 0.18 mmol) and sodium iodide (0.9 mmol) in dimethylformamide (2 mL) under nitrogen was heated at 60° C. for 3 hours. The reaction was then poured into water and extracted with diethyl ether. Drying over magnesium sulphate, concentration under reduced pressure and purification by flash chromatography followed by recrystallised from methanol gave the title compound (40 mg).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.80 (s, 3H), 0.82 (s, 3H), 0.95 (d, 3H), 1.01 (a, 3H), 1.02 (s, 3H), 3.10-3.31 (m, 2H), 5.35 (s, 1H). The mass spectrum showed characteristic peaks at: 496.1 (M⁺).

EXAMPLE 36 4,4-Dimethyl-17β-((1R)-methylbutyl)androsta-8,14-dien-3β-ol

To a solution of 24-toluenesulphonyloxy-4,4-dimethyl-5α-chola-8,14-diene-3-ol (100 mg, 0.18 mmol) in diethyl ether (25 mL) was added lithium aluminium hydride (40 mg, 1.1 mmol) and the reaction was stirred 16 hours at room temperature. The reaction was then poured into water and extracted with diethyl ether. Drying over magnesium sulphate, concentration under reduced pressure and purification by flash chromatography gave the title compound (41 mg).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.80 (s, 3H), 0.82 (s, 3H), 0.87 (t, 3H), 0.95 (d, 3H), 1.00 (s, 3H), 1.02 (s, 3H), 3.20-3.31 (m, 1H), 5.35 (m, 1H). The ¹³C-NMR spectrum (CDCl₃, d) showed characteristic signals at: 79.1, 117.8 (C-15), 123.2 (C-14), 142.1 (C-9), 151.4 (C-8). The mass spectrum showed characteristic peaks at: 370.3 (M⁺).

EXAMPLE 37 4,4-Dimethyl-17β-((1R)-methyl-4-cyanobutyl) androsta-8,14-dien-3β-ol

A mixture of 24-toluenesulphonylloxy-4,4-dimethyl-5α-chola-8,14-diene-3-ol (500 mg, 0.5 mmol) and sodium cyanide (90 mg, 2 mmol) in dimethylsulfoxide (2 mL) under nitrogen was heated at 140° C. for 2.5 hours. The reaction was then poured into ammonium chloride solution and extracted with dichloromethane. Drying over magnesium sulphate, concentration under reduced pressure and purification by flash chromatography gave the title compound (240 mg).

The ¹H-NMR spectrum (CDCl₃, d) showed characteristic signals at: 0.81 (s, 3H), 0.83 (s, 3H), 0.97 (d, 3H), 1.00 (s, 3H), 1.02 (s, 3H), 3.18-3.31 (m, 1H), 5.35 (m, 1H). The mass spectrum showed characteristic peaks at: 395.3 (M⁺).

EXAMPLE 38 27-Nor-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26-oic Acid Benzyl Ester

The compound is synthesised following the procedures outlined in example 52, below.

¹H-NMR (CDCl₃, 400 MHz): δ=7.35 (5H, m); 5.34 (1H, s); 5.11 (2H, s); 3.23 (1H, m).

EXAMPLE 39 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-(methionine Methyl Ester)amide

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl ester (18.0 g) is suspended in 300 ml of DMF and 14.8 g of imidazol, and 13 g of tert-butyldimethylsilylchloride is added. The reaction mixture is stirred at 70° C. for 20 hours. After aqueous work-up, and crystallisation from diethyl ether/methanol, 21.0 g of 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl ester is isolated. Melting point, 124-125° C.

¹H-NMR (CDCl₃, 400 MHz): δ=5.34 (1H, s); 3.67 (3H, s); 3.19 (1H, m); 0.9 (9H, s); 0.03 (6H, m).

3β-tert-Butyldimethylsilyfoxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl ester (3.6 g) is dissolved in a mixture of 150 ml of THF, 120 ml of ethanol and 18 ml of 1M sodium hydroxide. The mixture is stirred for 20 hours at room temperature and 2 hours at 50° C. After aqueous work-up and crystallisation from ethanol/water, 2.48 g of 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid is isolated.

¹H-NMR (CDCl₃, 300 MHz): δ=5.33 (1H, s); 3.19 (1H, m); 0.9 (9H, s); 0.03 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (0.5 g) is dissolved in 10 ml of dry dichloromethane and 0.213 ml of N-methylmorpholine. After cooling to ˜15° C. 0.132 ml of isobutylchloroformate is added and the mixture is stirred at −15° C. for 20 minutes, whereupon 0.232 g of L-methioninemethyl ester, hydrochloride is added and the mixture is stirred overnight and the temperature is slowly elevated to room temperature. After aqueous work-up, 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-(methionine methyl ester)amide (0.60 g) is obtained.

¹H-NMR (CDCl₃, 300 MHz): δ=6.12 (1H, d); 5.34 (1H, s); 4.73 (1H, m); 3.76 (3H, s); 3.2 (1H, m); 2.1 (3H, s); 9.0 (9H, s); 0.03 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-(methionine methyl ester)amide (0.20 g) is suspended in 20 ml methanol, and 0.1 ml 6M hydrogen chloride is added and the mixture is stirred at 50° C. for 1.5 hours and at room temperature for 20 hours. After crystallisation by adding of water, the title compound (165 mg) is obtained. Melting point: 138-141° C.

¹H-NMR (CDCl₃, 300 MHz): δ=6.12 (1H, d); 5.35 (1H, s); 4.73 (1H, m); 3.76 (3H, s); 3.23 (1H, m); 2.1 (3H, s). MS: Calculated: 545.8. Found 545.3.

EXAMPLE 40 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-(methionine)amide

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-(methionine methyl ester)amide (76 mg) is saponified overnight in a mixture of 10 ml of methanol, 5 ml of THF, 1 ml of water, and 0.6 ml of 1M sodium hydroxide. After aqueous work-up and crystallisation from methanol, the title compound (56 mg) is obtained.

¹H-NMR (CDCl₃, 300 MHz): δ=6.35 (1H, d); 5.35 (1H, s); 4.7 (1H, m); 3.25 (1H, m); 2.12 (3H, s). MS: Calculated: 531.8. Found 531.8.

EXAMPLE 41 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-(4-methylpiperazinyl)amide

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (0.40 g) is reacted with N-methylpiperazine and hydrolysed with HCl/ethanol following the procedure outlined in example 39 to give the title compound (80 mg). Melting point: 189-191° C.

¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 3.63 (2H, m); 3.48 (2H, m); 3.24 (1H, m); 2.31 (3H, s). MS: Calculated: 482.8. Found 482.3.

EXAMPLE 42 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-tert-butylamide

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (0.50 g) is reacted with tert-butylamine and hydrolysed with HCl/ethanol following the procedure outlined in example 39 to give the title compound (204 mg). Melting point: 171-176° C.

¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 5.21 (1H, s); 3.24 (1H, m); 1.36 (9H, s); MS: Calculated: 455.7. Found 455.4.

EXAMPLE 43 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-(isonipecotic Acid Ethyl Ester)amide

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (0.50 g) is reacted with ethylisonipecotate and hydrolysed with HCl/ethanol following the procedure outlined in example 39 to give the title compound (85 mg). Melting point: 116-119° C.

¹H-NMR (CDCl₃, 400 MHz): δ=5.35 (1H, s); 4.43 (1H, m); 4.15 (2H, q); 3.82 (1H, m); 3.25 (1H, m); 3.11 (1H, m); 2.8 (1H, m); 1.28 (3H, t). MS: Calculated: 539.8. Found 539.4.

EXAMPLE 44 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-(isonipecotic Acid)amide

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid-N-(isonipecotic acid ethyl ester)amide (36 mg) is saponified overnight in a mixture of 3 ml of ethanol and 0.2 ml of 1M sodiumhydroxide. After aqueous work-up and crystallisation from ethanol/water, the title compound (10 mg) is obtained. Melting point: 228-231° C.

¹H-NMR (CDCl₃, 400 MHz): δ=5.36 (1H, s); 4.45 (1H, m); 3.84 (1H, m); 3.27 (1H, m); 3.16 (1H, m); 2.86 (1H, m). MS: Calculated: 497.7. Found 497.6.

EXAMPLE 45 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid-N-(phenylalanine Methyl Ester)amide

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (0.40 g) is reacted with phenylalanine methyl ester and hydrolysed with HCl/methanol following the procedures outlined in example 39 to give the title compound (86 mg). Melting point: 158-160° C.

¹H-NMR (CDCl₃, 400 MHz): δ=7.27 (3H, m); 7.09 (2H, m); 5.86 (1H, d); 5.35 (1H, s); 4.89 (1H, m); 3.73 (3H, s); 3.25 (1H, m); 3.13 (2H, m). MS: Calculated: 561.8. Found 561.5.

EXAMPLE 46 3β-Hydroxy-4,4-dimethylchola-5,7-dien-24-oic Acid

3β-Hydroxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl ester (120 mg) is saponified in a mixture of 15 ml of methanol, 15 ml of THF and 0.7 ml of 1M sodium hydroxide at 50° C. After aqueous work-up and crystallisation from methanol, the title compound (7.6 mg) is isolated. Melting point: 210-213° C.

¹H-NMR (CDCl₃, 400 MHz): δ=5.92 (1H, d); 5.55 (1H, m); 3.38 (1H, m). MS: Calculated: 400.6. Found 400.2.

EXAMPLE 47 3β-Hydroxy-4,4-dimethylchola-5,7-dien-24-oic Acid-N-dimethyl Amide

3β-tert-Butyldimethylsilyloxy-4,4-dimethylchola-5,7-dien-24-oic acid methyl ester (3.5 g) is saponified and reacted with dimethylamine following the procedure outlined in example 39. The tert-butyldimethylsilyl protecting group is split of by treatment with tetra-butylammoniumfluoride hydrate according to the procedure outlined in example 2 to give the title compound.

¹H-NMR (CDCl₃, 400 MHz): δ=5.92 (1H, d); 5.54 (1H, m); 3.4 (1H, m); 3.03 (3H, m); 2.96 (3H, s). MS: Calculated: 427.7. Found 427.4.

EXAMPLE 48 4,4-Dimethyl-24-acetamido-5α-chola-8,14-dien-3β-ol

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-cic acid (5.0 g) is reacted with ammonia following the procedure outlined in example 39 and reduced with lithium aluminium hydride (3.0 g) in THF at room temperature. Aqueous work-up and crystallisation from methanol gives 2.6 g of 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-24-amino-5α-chola-8,14-diene.

¹H-NMR (CDCl₃, 300 MHz): δ=5.34 (1H, s); 3.2 (1H, m); 3.0 (2H, broad s); 2.7 (1H, m); 0.9 (9H, s); 0.03 (6H, m). MS: Calculated: 499.9. Found 499.4.

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-24-amino-5α-chola-8,14-diene (0.50 g) is acetylated in a mixture of 20 ml of pyridine and 7 ml of acetic anhydride and hydrolysed with HCl/ethanol. After aqueous work-up and crystallisation from ethanol/water, the title compound (0.24 g) is isolated. Melting point: 219-221° C.

¹H-NMR (CDCl₃, 300 MHz): δ=5.43 (1H, s); 5.35 (1H, s); 3.23 (3H, m); 1.98 (3H, s). MS: Calculated: 427.7. Found 427.4.

EXAMPLE 49 4,4-Dimethyl-24-acetoxy-5α-chola-8,14-dien-3β-ol

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid methyl ester (5.29 g) is reduced with lithium aluminium hydride (1.75 g) in 300 ml of THF at room temperature. After aqueous work-up and crystallisation from ethanol/water, 4.58 g of 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-ol is isolated.

¹H-NMR (CDCl₃, 400 MHz): δ=5.35 (1H, s); 3.65 (2H, m); 3.2 (1H, m); 0.9 (9H, s); 0.03 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-ol (150 mg) is acetylated in a mixture of 2 ml of pyridine and 1 ml of acetic anhydride. The tert-butyldimethylsilyl protecting group is split of by treatment with tetra-butylammonium fluoride hydrate according to the procedure outlined in example 2. After column chromatography and crystallisation from acetone/water, the title compound (36 mg) is obtained. ¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 4.05 (2H, m); 3.23 (1H, m); 2.04 (3H, s). MS: Calculated: 428.7. Found 428.3.

EXAMPLE 50 4,4-Dimethyl-24-methoxy-5α-chola-8,14-dien-3β-ol

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-ol (100 mg) is methylated with sodium hydride 60% (16 mg) and methyl iodine (0.125 ml) in 1 ml of DMF. The protecting tert-butyldimethylsilyl group is split of by treatment with HCl/ethanol. After aqueous work-up and crystallisation from methanol/water, the title compound (5 mg) is isolated.

¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 3.36 (2H, m); 3.32 (3H, s); 3.23 (1H, m). MS: Calculated: 400.7. Found 400.3.

EXAMPLE 51 4,4-Dimethyl-24-benzyloxy-5α-chola-8,14-dien-3β-ol

The compound is synthesised following the procedure in example 50. Melting point: 114-115° C.

¹H-NMR (CDCl₃, 300 MHz): δ=7.38-7.23 (5H, m); 5.35 (1H, s); 4.51 (2H, s); 3.45 (2H, m); 3.24 (1H, m). MS: Calculated: 476.7. Found 476.3.

EXAMPLE 52 3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic Acid Benzyl Ester

3β-Hydroxy-4,4-dimethyl-5α-chola-8,14-dien-24-oic acid (100 mg) is suspended in 5 ml of dry DMF. 811 mg of cesium carbonate and 0.29 ml of benzyl chloride is added and the mixture is stirred at 50° C. overnight. After aqueous work-up, column chromatography and crystallisation from acetone/water 55 mg of the title compound is obtained. Melting point: 118-119° C.

¹H-NMR (CDCl₃, 300 MHz): δ=7.35 (5H, m); 5.35 (1H, s); 5.12 (2H, s); 3.23 (1H, m). MS: Calculated: 490.7. Found 490.3.

EXAMPLE 53 26,27-Diethyl-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioate

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-5α-chola-8,14-dien-24-ol (4.0 g) is dissolved in 80 ml of dry pyridine at 0° C. 3.04 g of p-toluenesulphonyl chloride is added and the mixture is stirred at 5° C. for 20 hours and at room temperature for 5 hours. After aqueous work-up and crystallisation from methanol, 3.01 g of 3β-tert-butyldimethylsilyloxy-4,4-dimethyl-24-p-toluenesulphonyloxy-5α-chola-8,14-diene is obtained.

¹H-NMR (CDCl₃, 300 MHz): δ=7.79 (2H, d); 7.34 (2H, d); 5.33 (1H, s); 4.02 (2H, m); 3.19 (1H, m); 2.43 (3H, s); 0.9 (9H, s); 0.03 (6H, m).

3β-tert-Butyldimethylsilyloxy-4,4-dimethyl-24-p-toluenesulphonyloxy-5α-chola-8,14-diene (0.50 g) is added to a mixture of 0.58 ml of diethyl malonate and 137 mg of sodium hydride (60%) in 10 ml of dry THF at −70° C. Stirring at −70° C. for 15 minutes, whereupon elevating of the temperature and reflux for 9 hours. After aqueous work-up and crystallisation from ethanol/water, 0.432 g of 26,27-diethyl-3β-tertbutyldimethylsilyloxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioate is obtained.

¹H-NMR (CDCl₃, 300 MHz): δ=5.33 (1H, s); 4.21 (4H, q); 3.32 (1H, t); 3.2 (1H, m); 0.9 (9H, s); 0.03 (6H, m). MS: Calculated: 643.0. Found 642.4.

26,27-Diethyl-3β-tert-butyldimethylsilyloxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioate (0.41 g) is dissolved in 40 ml of ethanol and 0.5 ml of 6M hydrogen chloride. Stirring at room temperature for 48 h and 40° C. for 1 hour. The product is crystallised by adding of water and recrystallised from ethanol to give. 141 mg of the title compound. Melting point: 105-106° C.

¹H-NMR (CDCl₃, 300 MHz): δ=5.35 (1H, s); 4.2 (4H, q); 3.33 (1H, t); 3.25 (1H, m); 1.25 (6H, t). MS: Calculated: 528.8. Found 528.4.

EXAMPLE 54 3β-Hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioic Acid

26,27-Diethyl-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioate (95 mg) is saponified in a mixture of 5 ml of 96% ethanol, 5 ml of THF and 5 ml of 1M sodium hydroxide at room temperature. After aqueous work-up, the title compound is isolated (60 mg). Melting point: 188-190° C.

¹H-NMR (DMSO-d₆, 400 MHz): δ=12.63 (2H, s); 5.3 (1H, s); 4.37 (1H, s); 3.2 (1H, t); 3.03 (1H, m). MS: Calculated: 472.7. Found 473.2 [M+H]⁺.

EXAMPLE 55 27-Nor-3β-hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26-oic Acid

3β-Hydroxy-4,4-dimethyl-5α-cholesta-8,14-dien-26,27-dioic acid (535 mg) is suspended in 30 ml of o-xylene and refluxed overnight. After evaporation of o-xylene, the title compound is isolated by crystallisation from methanol/ether. Yield: 338 mg.

¹H-NMR (DMSO-d₆, 400 MHz): δ=11.95 (1H, s); 5.28 (1H, s); 4.35 (1H, m); 3.0 (1H, m). MS: Calculated: 428.7. Found 428.2.

EXAMPLE 56 Synthesis of the Intermediate Cholesta-4,8-dien-3-one

A solution of 2.20 g cholesta-5,8-dien-3β-ol in 27 ml toluene and 6 ml cyclohexanone is refluxed for 10 minutes in a Dean-Stark apparatus. 0.57 g aluminum isopropoxide are added and the reaction mixture is refluxed for 30 minutes. After cooling and addition of sulfuric acid (2 N), the resulting mixture is extracted with ethyl acetate. The organic layer is separated, washed with saturated sodium bicarbonate solution and water, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of hexane and ethyl acetate to give 1.53 g cholesta-4,8-dien-3-one as a white solid.

¹H-NMR (CDCl₃): δ=0.68 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.94 (d, J=7 Hz, 3H, H-21); 1.35 (s, 3H, H-19); 5.77 (s, 1H, H-4).

Cholesta-4,8(14)-dien-3-one is synthesised from cholesta-5,8(14)-dien-3β-ol in the same way. Cholesta-4,7-dien-3-one is synthesised according to literature procedures [Just. Liebigs Ann. Chem. 542 (1939), 218,].

Example 57 Scheme 1 Cholesta-4,8-dien-3α-ol and Cholesta-4,8-dien-3β-ol

To a solution of 520 mg cholesta-4,8-dien-3-one in 45 ml tetrahydrofuran 1.70 ml of a L-Selectride solution (1 N, in tetrahydrofuran) are added dropwise at −75° C. The reaction mixture is warmed to room temperature within 4 hours, poured into hydrochloric acid (1 N) and extracted with ethyl acetate. The organic layer is separated, washed with brine, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of hexane and ethyl acetate to give 286 mg cholesta-4,8-dien-3β-ol and 20 mg cholesta-4,8-dien-3α-ol as white solids.

Cholesta-4,8-dien-3α-ol

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.86 (2×d, J=7 Hz, 6H, H-26/27); 0.92 (d, J=7 Hz, 3H, H-21); 1.11 (s, 3H, H-19); 4.04 (m, 1H, H-3); 5.47 (d, H=5 Hz, 1H, H-4).

Cholesta-4,8-dien-3β-ol

¹H-NMR (CDCl₃): δ=0.64 (s, 3H, H-18); 0.86 (2×d, J=7 Hz, 6H, H-26/27); 0.93 (d, J=7 Hz, 3H, H-21); 1.23 (s, 3H, H-19); 2.47 (m, 1H); 4.19 (m, 1H, H-3); 5.32 (s, 1H, H-4).

Cholesta-4,7-dien-3α-ol and cholesta-4,7-dien-3β-ol are synthesised according to literature procedures [Just. Liebigs. Ann. Chem. 542 (1939), 218].

EXAMPLE 58 Scheme 2 a) 5-Cyano-5β-cholest-8-en-3-one and 5-Cyano-5α-cholest-8-en-3-one

30 ml of a diethylaluminum cyanide solution (1 N, in toluene) are added to a solution of 3.82 g cholesta-4,8-dien-3-one in 60 ml tetrahydrofuran at 0° C. The reaction mixture is warmed to room temperature and stirred for 4 hours. 20 ml of a sodium hydroxide solution (1 N) are added before the mixture is extracted with diethyl ether. The organic layer is separated, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of hexane and ethyl acetate to give 1.46 g 5-cyano-5β-cholest-8-en-3-one and 1.76 g 5-cyano-5α-cholest-8-en-3-one as pale yellow crystals.

5-Cyano-5α-cholest-8-en-3-one

¹H-NMR (CDCl₃): δ=0.69 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.95 (d, J=7 Hz, 3H, H-21); 1.43 (s, 3H, H-19); 2.63 (m, 1H).

5-Cyano-5α-cholest-8-en-3-one

¹H-NMR (CDCl₃): δ=0.64 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.95 (d, J=7 Hz, 3H, H-21); 1.37 (s, 3H, H-19); 2.52 (m, 2H).

5-Cyano-5β-cholest-7-en-3-one [which is known from the literature: Aust. J. Chem. 35 (1982), 629], 5-cyano-5α-cholest-7-en-3-one, 5-cyano-5β-cholest-8(14)-en-3-one and 5-cyano-5α-cholest-8(14)-en-3-one are synthesised from the corresponding starting materials in the same way.

b) 5-Cyano-5α-cholest-8-en-3β-ol and 5-Cyano-5α-cholest-8-en-3α-ol

327 mg sodium borohydride are added to a solution of 1.76 g 5-cyano-5α-cholest-8-en-3-one in 200 ml ethanol at room temperature. The reaction mixture is stirred for 4 hours. After addition of hydrochloric acid (1 N) the resulting mixture is extracted with dichloromethane. The organic layer is separated, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of hexane and ethyl acetate to give 0.36 mg 5-cyano-5α-cholest-8-en-3β-ol and 1.00 g 5-cyano-5α-cholest-8-en-3α-ol as white solids.

5-Cyano-5α-cholest-8-en-3β-ol

¹H-NMR (CDCl₃): δ=0.59 (s, 3H, H-18); 0.86 (2×d, J=7 Hz, 6H, H-26/27); 0.92 (d, J=7 Hz, 3H, H-21); 1.08 (s, 3H, H-19); 4.12 (bm, 1H, H-3).

5-Cyano-5α-cholest-8-en-3α-ol

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.89 (2×d, J=7 Hz, 6H, H-26/27); 0.94 (d, J=7 Hz, 3H, H-21); 1.06 (s, 3H, H-19); 4.11 (bm, 1H, H-3).

5-Cyano-5α-cholest-7-en-3β-ol, 5-cyano-5α-cholest-7-en-3α-ol, 5-cyano-5β-cholest-7-en-3α-ol, 5-cyano-5β-cholest-8-en-3β-ol, 5-cyano-5β-cholest-8-en-3α-ol, 5-cyano-5α-cholest-8(14)-en-3β-ol, 5-cyano-5α-cholest-8(14)-en-3α-ol, 5-cyano-5β-cholest-8(14)-en-3β-ol 5-cyano-5β-cholest-8(14)-en-3α-ol are synthesised in the same way.

EXAMPLE 59 Scheme 3 a) 3β-Hydroxy-5α-cholest-8-ene-5-carbaldehyde

4.66 ml of a diisobutylaluminum hydride solution (1.2 N; in toluene) are added to a solution of 230 mg 5-cyano-5α-cholest-8-en-3β-ol in 18 ml toluene at −1° C. The reaction mixture is stirred for 3 hours before 3.6 ml sulfuric acid (1 N) are added. After refluxing for one hour the resulting mixture is cooled to room temperature, diluted with water and extracted with dichloromethane. The organic layer is separated, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent 220 mg 3β-hydroxy-5αcholest-8-ene-5-carbaldehyde are obtained as a white solid.

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.86 (2×d, J=7 Hz, 6H, H-26/27); 0.94 (d, J=7 Hz, 3H, H-21); 1.17 (s, 3H, H-19); 2.23 (m, 2H); 3.61 (m, 1H, H-3); 9.86 (s, 1H, 5-CHO).

b) 5-(Hydroxymethyl)-5α-cholest-8-en-3β-ol

A suspension of 56 mg lithiumaluminum hydride in 5 ml tetrahydrofuran is added to a solution of 200 mg 3β-hydroxy-5α-cholest-8-en-5-carbaldehyde in 20 ml tetrahydrofuran at room temperature. After being heated to 50° C. for two hours the reaction mixture is cooled to room temperature, 0.06 ml water, 0.06 ml of a sodium hydroxide solution (1 N) and 0.18 ml water are added subsequently. The resulting suspension is stirred for 15 minutes and filtered over anhydrous sodium sulphate. After evaporation of the solvent the residue is crystallized from ethylacetate to give 80 mg 5-(hydroxymethyl)-5α-cholest-8-en-3β-ol as a white solid.

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.93 (d, J=7 Hz, 3H, H-21); 1.16 (s, 3H, H-19); 3.58 (s, 2H, 5-CH₂OH); 3.97 (m, 1H, H-3).

5-(Hydroxymethyl)-5α-cholest-7-en-3β-ol, 5-(hydroxymethyl)-5β-cholest-8-en-3α-ol and 5-(hydroxymethyl)-5α-cholest-8(14)-en-3β-ol are synthesised in the same way.

EXAMPLE 60 Scheme 4 3′,4α-Dihydrocyclopropa[4,5]-5β-cholest-8-en-3β-ol

300 mg zinc dust and 0.03 ml glacial acetic acid are added to a solution 5.4 mg cupric acetate in 1.2 ml dimetoxyethane at room temperature. The mixture is stirred for 30 minutes before 0.01 ml triethylamine are added. After 5 minutes a solution of 100 mg cholesta-4,8-dien-3b-ol in 0.4 ml, dimethoxyethane is added. Then 0.24 ml diiodomethane are added at such a rate that the reaction temperature does not rise above 40° C. The reaction mixture is stirred for further 6 hours. After addition of saturated ammonium chloride solution the resulting mixture is extracted with ethyl acetate. The organic layer is separated, washed with brine, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of hexane and ethyl acetate to give 32 mg.

¹H-NMR (CDCl₃): δ=0.25 (dd, J=9 Hz, 5 Hz, 1H, 4,5-CH₂); 0.63 (s, 3H, H-18); 0.76 (dd, J=5 Hz, 5 Hz, 1H, 4,5-CH₂); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.94 (d, J=7 Hz, 3H, H-21); 1.11 (s, 3H, H-19); 4.32 (m, 1H, H-3).

EXAMPLE 61 Scheme 5 a) 5-Methyl-5β-cholest-8-en-3-one

0.85 ml of a methyllithium solution (1.6 N, in diethyl ether) are added to a suspension of 130 mg cuprous iodide in diethyl ether at 0° C. The resulting mixture is stirred for one hour before a solution of 130 mg cholesta-4,8-dien-3-one in 1 ml diethyl ether is added. After being stirred for 30 minutes the reaction mixture is poured into saturated ammonium chloride solution and extracted with ethyl acetate. The organic layer is separated, washed with brine, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with mixture of hexane and ethyl acetate to give 80 mg 5-methyl-5β-cholest-8-en-3-one as a white solid.

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.86 (2×d, J=7 Hz, 6H, H-26/27); 0.90 (s, 3H, 5-CH₃); 0.93 (d, J=7 Hz, 3H, H-21); 1.06 (s, 3H, H-19); 2.42 (m, 1H).

b) 5-Methyl-5β-cholest-8-en-3α-ol and 5-Methyl-5β-cholest-8-en-3β-ol

0.31 ml of a K-Selectride solution (1 N, in tetrahydrofuran) are added to a solution of 63 mg 5-methyl-5β-cholest-8-en-3-one in 4.4 ml tetrahydrofuran at −65° C. After being stirred for 2 hours the reaction mixture is allowed to warm to room temperature, poured into saturated ammonium chloride solution and extracted with ethyl acetate. The organic layer is separated, washed with brine, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of dichloromethane and acetone to give 26 mg 5-methyl-5β-cholest-8-en-3α-ol and 21 mg 5-methyl-5β-cholest-8-en-3β-ol as white solids.

5-Methyl-5β-cholest-8-en-3α-ol

¹H-NMR (CDCl₃): δ=0.59 (s, 3H, H-18); 0.85 (2×d, J=7 Hz, 6H, H-26/27); 0.86 (s, 3H); 0.90 (s, 3H); 0.92 (d, J=7 Hz, 3H, H-21); 3.82 (m, 1H, H-3).

5-Methyl-5β-cholest-8-en-3β-ol

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.89 (s, 3H, 5-CH₃); 0.93 (d, J=7 Hz, 3H, H-21); 0.99 (s, 3H, H-19); 3.87 (m, 1H, H-3).

5-Methyl-5β-cholest-8(14)-en-3-one, 5-methyl-5β-cholest-8(14)-en-3β-ol and 5-methyl-5β-cholest-8(14)-en-3α-ol are synthesised in the same way.

EXAMPLE 62 Scheme 1 3β-(Trifluoromethyl)cholesta-4,8-dien-3α-ol and 3α-(Trifluoromethyl)cholesta-4,8-dien-3β-ol

0.2 ml triethylsilyltrifuoromethane and 328 mg tetrabutylammonium fluoride trihydrate are added to a solution of 200 mg cholesta-4,8-dien-3-one in 10 ml tetrahydrofuran at room temperature. After being stirred for 2 hours the reaction mixture is diluted with ethyl acetate, washed with water and brine, dried over anhydrous sodium sulphate and filtered. After evaporation of the solvent the residue is chromatographed with a mixture of hexane and ethyl acetate to give 45 mg 3β-(trifluoromethyl)cholesta-4,8-dien-3α-ol as a colourless oil and 170 mg 3α-(trifluoromethyl)cholesta-4,8-dien-3β-ol as a white solid.

3β-(Trifluoromethyl)cholesta-4,8-dien-3α-ol

¹H-NMR (CDCl₃): δ=0.65 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.93 (d, J=7 Hz, 3H, H-21); 1.18 (s, 3H, H-19); 2.41 (m, 1H,); 5.39 (s, 1H, H-4).

3α-(Trifluoromethyl)cholesta-4,8-dien-3β-ol

¹H-NMR (CDCl₃): δ=0.64 (s, 3H, H-18); 0.87 (2×d, J=7 Hz, 6H, H-26/27); 0.93 (d, J=7 Hz, 3H, H-21); 1.25 (s, 3H, H-19); 2.43 (m, 1H,); 5.25 (s, 1H, H-4).

EXAMPLES 63+64 (20R)-5-Cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3β-ol and (20R)-5-Cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3α-ol a) Ergosta-4,8,22-trien-3-one

1.90 g ergosta-5,8,22-trien-3β-ol (lichesterol) were treated with 0.50 g aluminum-tris-iso-propylate as described in example 56. Column chromatography gave 1.54 g ergosta-4,8,22-trien-3-one as a white solid.

¹H-NMR (CDCl₃): δ=0.69 (s, 3H, H-18); 0.82-1.05 (4×d, 4×Me); 1.36 (s, 3H, H-₁); 5.22 (m, 2H, H-22/23); 5.77 (s, 1H, H-4).

b) 5-Cyano-5α-ergosta-8,22-dien-3-one

1.54 g ergosta-4,8,22-trien-3-one were treated with 11.73 ml diethylaluminum cyanide solution (1 N, toluene) as described in example 58. After aqueous work up and chromatography 0.90 g 5-cyano-5α-ergosta-8,22-dien-3-one were isolated as a white solid (beside the corresponding 5β-cyano-compound).

¹H-NMR (CDCl₃): δ=0.66 (s, 3H, H-18); 0.82-1.05 (4×d, 4×Me); 1.28 (s, 3H, H-19); 5.21 (m, 2H, H-22/23).

c) 5-Cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-ergosta-8,22-diene

A mixture of 900 mg 5-cyano-5α-ergosta-8,22-dien-3-one, 0.97 ml ethylene glycol, 25 mg p-toluenesulfonic acid in 20 ml toluene was refluxed at a dean-stark-trap for 2 hours. After cooling, the reaction mixture was poured into saturated sodium bicarbonate solution, extracted with ethyl acetate and washed with water. The combined organic extracts were dried and evaporated to give 900 mg 5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-ergosta-8,22-diene as a white solid.

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.82-1.05 (4×d, 4×Me); 1.11 (s, 3H, H-19); 3.90-4.15 (m, 4H, 3-ketal); 5.20 (m, 2H, H-22/23).

d) (20R)-5-Cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol

450 mg 5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-ergosta-8,22-diene were treated with ozone as described in example 19b. After reductive work up, 172 mg (20R)-5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol were isolated as a white solid.

¹H-NMR (CDCl₃): δ=0.65 (s, 3H, H-18); 1.06 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 3.35 (m, 1H, H-22); 3.66 (m, 1H, H-22); 3.904.12 (m, 4H, 3-ketal).

e) (20R)-5-Cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol-4-methylbenzene Sulfonate

1.13 g (20R)-5-cyano-5α-pregn-8-ene-20-methanol was treated with 869 mg p-toluenesulfonyl chloride as described in example 19d. After aqueous work up and column chromatography, 1.19 g (20R)-5-cyano-3-(spiro-2′,5′-dioxacyclopentyl)-5α-pregn-8-ene-20-methanol-4-methylbenzene sulfonate were isolated.

¹H-NMR (CDCl₃): δ=0.59 (s, 3H, H-18); 1.00 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 3.70-4.12 [m, 4H (3-ketal)+2H (H-22)].

f) (20R)-5-Cyano-21-cyclohexyl-20-methyl-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene

258 mg (20R)-5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol-4-methylbenzene sulfonate were treated with cyclohexyl magnesium bromide analoguously to example 19e. After column chromatography, 147 mg (20R)-5-cyano-21-cyanohexyl-20-methyl-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene were isolated as a pale yellow solid.

¹H-NMR (CDCl₃): δ=0.62 (s, 3H, H-18); 0.90 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 3.90-4.12 (m, 4H, 3-ketal).

g) (20R)-5-Cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3-one

A mixture of 128 mg (20R)-5-cyano-21-cyclohexyl-20-methyl-3-(spiro-2′,5′-dioxacyclopentyl)-5α-pregn-8-ene, 31 mg amberlyste 15 and 8 ml acetone was stirred at room temperature for 20 hours. After filtration and evaporation of the solvent 94 mg (20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3-one were isolated.

¹H-NMR (CDCl₃): δ=0.67 (s, 3H, H-18); 0.90 (d, J=7 Hz, 3H, H-21); 1.27 (s, 3H, H-19); 2.55 (pseudo-s, 2H).

h) (20R)-5-Cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3β-ol and (20R)-5-Cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3α-ol

90 mg (20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3-one were treated with 33 mg sodium borohydride as described in example 58b. After column chromatography, 15 mg (20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3β-ol and 25 mg (20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3α-ol were isolated.

(20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3β-ol

¹H-NMR (CDCl₃): δ=0.63 (s, 3H, H-18); 0.93 (d, J=7 Hz, 3H, H-21); 1.11 (s, 3H, H-19); 4.12 (broad-m, 1H, H-3).

(20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3α-ol

¹H-NMR (CDCl₃): δ=0.62 (s, 3H, H-18); 0.92 (d, J=7 Hz, 3H, H-21); 1.06 (s, 3H, H-19); 4.12 (narrow-m, 1H, H-3).

EXAMPLES 65+66 (20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3β-ol and (20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3α-ol a) (20R)-5-Cyano-21-phenyl-20-methyl-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene

400 mg (20R)-5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol-4-methylbenzene sulfonate (example 63e) were treated with phenyl magnesium bromide analoguously to example 63f. After column chromatography, 190 mg (20R)-5-cyano-21-phenyl-20-methyl-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene were isolated as a white solid.

¹H-NMR (CDCl₃): δ=0.65 (s, 3H, H-18); 0.84 (d, J=7 Hz, 3H, H-21); 1.12 (s, 3H, H-19); 2.90 (dd, J=12 hz, J=3 Hz, 1H, H-21); 3.90-4.12 (m, 4H, 3-ketal); 7.12-7.30 (m, 5H, ph).

b) (20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3-one

180 mg (20R)-5-cyano-21-phenyl-20-methyl-3(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene were treated with-amberlyste 15 as described in example 63g. After filtration and evaporation of the solvent, 164 mg (20R)-5-cyano-21-phenyl-20-methyl-5α-pregn-8-en-3-one were isolated.

¹H-NMR (CDCl₃): δ=0.68 (s, 3H, H-18); 0.84 (d, J=7 Hz, 3H, H-21); 1.27 (s, 3H, H-19); 2.55 (pseudo-s, 2H); 2.90 (dd, J=12 hz, J=3 Hz, 1H, H-21); 7.12-7.30 (m, 5H, ph).

c) (20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3β-ol and (20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3α-ol

152 mg (20R)-5-cyano-21-cyclohexyl-20-methyl-5α-pregn-8-en-3-one were treated with 50 mg sodium borohydride as described in example 58b. After column chromatography 36 mg (20R)-5-cyano-21-phenyl-20-methyl-5α-pregn-8-en-3β-ol and 46 mg (20R)-5-cyano-21-phenyl-20-methyl-5α-pregn-8-en-3α-ol were isolated.

(20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3β-ol

¹H-NMR (CDCl₃): δ=0.67 (s, 3H, H-18); 0.85 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 2.90 (dd, J=12 hz, J=3 Hz, 1H, H-21); 4.15 (broad-m, 1H, H-3); 7.12-7.30 (m, 5H, ph).

(20R)-5-Cyano-21-phenyl-20-methyl-5α-pregn-8-en-3α-ol

¹H-NMR (CDCl₃): δ=0.65 (s, 3H, H-18); 0.84 (d, J=7 Hz, 3H, H-21); 1.05 (s, 3H, H-19); 2.90 (dd, J=12 hz, J=3 Hz, 1H, H-21); 4.12 (narrow-m, 1H, H-3); 7.12-7.30 (m, 5H, ph).

EXAMPLES 67+68 5-Cyano-24-nor-5α-cholest-8-en-3β-ol and 5-Cyano-24-nor-5α-cholest-8-en-3α-ol a) 5-Cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-24-nor-5α-cholest-8-ene

400 mg (20R)-5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol-4-methylbenzene sulfonate (example 63e) were treated with isobutyl magnesium bromide analoguously to example 63f. After column chromatography 206 mg 5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-24-nor-5α-cholest-8-ene were isolated as a white solid.

¹H-NMR (CDCl₃): δ=0.62 (s, 3H, H-18); 0.85-0.95 (3×d, J=7 Hz); 1.10 (s, 3H, H-19); 3.88-4.10 (m, 4H, 3-ketal).

b) 5-Cyano-24-nor-5α-cholest-8-en-3-one

188 mg 5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-24-nor-5α-cholest-8-ene were treated with amberlyste 15 as described in example 63g. After filtration and evaporation of the solvent, 175 mg 5-cyano-24-nor-5α-cholest-8-en-3-one were isolated.

¹H-NMR (CDCl₃): δ=0.65 (s, 3H, H-18); 0.85-0.95 (3×d, J=7 Hz); 1.27 (s, 3H, H-19); 2.55 (pseudo-s, 2H).

c) 5-Cyano-24-nor-5α-cholest-8-en-3β-ol and 5-Cyano-24-nor-5α-cholest-8-en-3α-ol

173 mg 5-cyano-24-nor-5α-cholest-8-en-3-one were treated with 67 mg sodium borohydride as described in example 58b. After column chromatography, 35 mg 5-cyano-24-nor-5α-cholest-8-en-3β-ol and 64 mg 5-cyano-24-nor-5α-cholest-8-en-3α-ol were isolated.

5-Cyano-24-nor-5α-cholest-8-en-3β-ol

¹H-NMR (CDCl₃): δ=0.60 (s, 3H, H-18); 0.85-0.95 (3×d, J=7 Hz); 1.10 (s, 3H, H-19); 4.13 (broad-m, 1H, H-3).

5-Cyano-24-nor-5α-cholest-8-en-3α-ol

¹H-NMR (CDCl₃): δ=0.62 (s, 3H, H-18); 0.85-0.95 (3×d, J=7 Hz); 1.05 (s, 3H, H-19); 4.10 (broad-m, 1H, H-3).

EXAMPLES 69+70 5-Cyano-24-nor-5α-cholesta-8,23-dien-3β-ol and 5-Cyano-24-nor-5α-cholesta-8,23-dien-3α-ol a) 5-Cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-24-nor-5α-cholesta-8,23-diene

400 mg (20R)-5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-5α-pregn-8-ene-20-methanol-4-methylbenzene sulfonate (example 63e) were treated with grignard reagent from 1-bromo-2-methylpropene analoguously to example 63f. After column chromatography, 206 mg 5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-24-nor-5α-cholesta-8,23-diene were isolated as a white solid.

¹H-NMR (CDCl₃): δ=0.62 (s, 3H, H-18); 0.92 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 1.59, 1.70 (2×s, 2×3H); 3.90-4.10 (m, 4H, 3-ketal); 5.12 (t, J=7 Hz; 1H, H-23).

b) 5-Cyano-24-nor-5α-cholesta-8,23-dien-3-one

190 mg 5-cyano-3-(spiro-2′,5′-dioxa-cyclopentyl)-24-nor-5α-cholesta-8,23-diene were treated with amberlyste 15 as described in example 63g. After filtration and evaporation of the solvent, 170 mg 5-cyano-24-nor-5α-cholesta-8,23-dien-3-one were isolated. ¹H-NMR (CDCl₃): δ=0.60 (s, 3H, H-18); 0.90 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 1.61, 1.72 (2×s, 2×3H); 2.55 (pseudo-s, 2H); 5.12 (t, J=7 Hz; 1H, H-23).

c) 5-Cyano-24-nor-5α-cholesta-8,23-dien-3β-ol and 5-Cyano-24-nor-5α-cholesta-8,23-dien-3α-ol

170 mg 5-cyano-24-nor-5α-cholesta-8,23-dien-3-one were treated with 67 mg sodium borohydride as described in example 58b. After column chromatography and HPLC for purification, 8 mg 5-cyano-24-nor-5α-cholesta-8,23-dien-3β-ol and 19 mg 5-cyano-24-nor-5α-cholesta-8,23-dien-3α-ol were isolated.

5-Cyano-24-nor-5α-cholesta-8,23-dien-3β-ol

¹H-NMR (CDCl₃): δ=0.60 (s, 3H, H-18); 0.90 (d, J=7 Hz, 3H, H-21); 1.10 (s, 3H, H-19); 1.59, 1.72 (2×s, 2×3H); 4.13 (broad-m, 1H, H-3); 5.12 (t, J=7 Hz; 1H, H-23).

5-Cyano-24-nor-5α-cholesta-8,23-dien-3α-ol

¹H-NMR (CDCl₃): δ=0.61 (s, 3H, H-18); 0.90 (d, J=7 Hz, 3H, H-21); 1.05 (s, 3H, H-19); 1.59, 1.72 (2×s, 2×3H); 4.11 (narrow-m, 1H, H-3); 5.12 (t, J=7 Hz; 1H, H-23).

EXAMPLE 71

An Agonistic Oocyte Assay Can Be Performed as Follows:

Oocytes were obtained from immature female mice (C57BL/6J×DBA/2J F1, Bomholtgaard, Denmark) weighing 13-16 grams, that were kept under controlled temperature (20-22° C.), light (lights on 06.00-18.00) and relative humidity (50-70%). The mice received an intra-peritoneal injection of 0.2 ml gonadotropins (Gonal-F, Serono) containing 20 IU FSH and 48 hours later the animals were killed by cervical dislocation.

The ovaries were dissected out and the oocytes were isolated in Hx-medium (see below) under a stereo microscope by manual rupture of the follicles using a pair of 27 gauge needles. Spherical oocytes displaying an intact germinal vesicle (hereinafter designated GV) were divided in cumulus enclosed oocytes (hereinafter designated CEO) and naked oocytes (hereinafter designated NO) and placed in α-minimum essential medium (α-MEM without ribonucleosides, Gibco BRL, Cat. No. 22561) supplemented with 3 mg/ml bovine serum albumin (BSA, Sigma Cat. No. A-7030), 5 mg/ml human serum albumin (HSA, Statens Seruminstitute, Denmark), 0.23 mM pyruvate (Sigma, Cat. No S-8636), 2 mM glutamine (Flow Cat. No. 16-801), 100 IU/ml penicillin and 100 μg/ml streptomycin (Flow, Cat No. 16-700). This medium was supplemented with 3 mM hypoxanthine (Sigma Cat. No. H-9377) and designated Hx-medium.

The oocytes were rinsed three times in Hx-medium and oocytes of uniform size were divided into groups of CEO and NO. CEO and NO were cultured in 4-well multidishes (Nunclon, Denmark) in which each well contained 0.4 ml of Hx-medium. One control well (i.e., 35-45 oocytes cultured in identical medium with no addition of test compound) was always cultured simultaneously with 3 test wells (35-45 oocytes per well supplemented with test compound).

The oocytes were cultured in a humidified atmosphere of 5% CO₂ in air for 24 hours at 37° C. By the end of the culture period, the number of oocytes with germinal vesicle (hereinafter designated GV), germinal vesicle breakdown (hereinafter designated GVB) and polar bodies (hereinafter designated PB), respectively, were counted using a stereomicroscope (Wildt, Leica MZ 12). The percentage GVB, defined as percentage of oocytes undergoing GVB per total number of oocytes in that well, was calculated as: %GVB=(number of GVB+number of PB/total number oocytes)×100.

The % PB was defined as percentage of oocytes displaying one extruded polar body per total number of oocytes in that well.

The effect of the tested compounds has been indexed against control level and 4,4-dimethyl-5α-cholesta-8,14,24-trien-3β-ol (hereinafter designated FF-MAS) where controls and FF-MAS are indexed to an effect of 0 and 100, respectively. The relative effect of the tested compound is calculated as follows: Relative effect=((test GVB %−control GVB %)/(FF-MAS GVB %−control GVB %))×100. Results

TABLE 1 The mean percentage GVB, the mean percentage PB and mean Relative Effect of compounds after culture of asked oocytes (NO) in vitro for 24 hours. Concentration mean mean mean mikroM % GVB % PB Relative Effect CONTROL 0 11.9 5.5 0 FF-MAS 10 86.1 28.5 100 Example 4 10 10.5 3.5 −10 Example 6 10 88 11 110 Example 10 10 10 3 −10 Example 21 10 82 29.5 98 Example 23 10 73.5 21.5 98 Example 33 10 70 14.5 105

EXAMPLE 72

An Antagonistic Oocyte Assay Can Be Performed as Follows:

Animals

Oocytes were obtained from immature female mice (C57BI/6J×DBA/2J F1-hybrids, Bomholtgaard, Denmark) weighing 13-16 grams, that were kept under controlled lighting and temperature. The mice received an intraperitoneal injection of 0.2 gonadotropins (Gonal F, Serono, Solna, Sweden, containing 20 IU FSH, alternatively, Puregon, Organon, Swords, Ireland containing 20 IU FSH) and 48 hours later the animals were killed by cervical dislocation.

Test of Meiosis-inhibiting Substances in the Oocyte Test

The ovaries were dissected out and the oocytes were isolated in Hx-medium (see below) under a stereo microscope by manual rupture of the follicles using a pair of 27 gauge needles. Spherical, naked oocytes (NO) displaying an intact germinalvesicle (GV) were placed in α-minimum essential medium (α-MEM without ribonucleosides, Gibco BRL, Cat.No. 22561) supplemented with 3 mM hypoxanthine (Sigma Cat. No. H-9377), 8 mg/ml human serum albumin (HSA, Statens Seruminstitut, Denmark), 0.23 mM pyrubate (Sigma, Cat. No. S-8636), 2 mM glutamine (Flow Cat. No. 16-801), 100 IU/ml penicillin and 100 μg/ml streptomycin (Flow, Cat No. 16-700). This medium was designated Hx-medium.

Naked oocytes (NO) were rinsed three times in Hx-medium. 4,4-Dimethyl-5α-cholesta-8,14,24-trien-3β-ol (FF-MAS) has previously been shown to induce meiosis in NO in vitro (Byskov, A. G. et al. Nature 374 (1995), 559-562). NO were cultured in Hx-medium supplemented with 5 μM FF-MAS in co-culture with the test compounds in different concentrations in 4-well multidishes (Nunclon, Denmark) in which each well contained 0.4 ml of the medium and 35-45 oocytes. One positive control (i.e., 35-45 oocytes cultured in Hx-medium containing FF-MAS with no addition of test compound) was always run simultaneously with the test cultures, which were supplemented with different concentrations of the compounds to be tested. In addition, one negative control (35-45 oocytes cultured in Hx-medium alone) was run simultaneously with the positive control.

Examination of Oocytes

By the end of the culture period, the number of oocytes with germinal vesicle (GV) or germinal vesicle breakdown (GVB) and those with polar body (PB) was counted using a stereomicroscope of an inverted microscope with differential interference contrast equipment. The percentage of oocytes with GVB+PB per total number of oocytes were calculated in the test cultures and in the control (positive and negative) culture groups. The relative inhibition of the test compound was calculated by the following formula: Inhibition of test compound (in percentage)=100−{(GBV_(test compound)−GBV_(negative control))×100/(GBV_(positive control)−GBV_(negative control))}.

In case of a dose response curve, an IC₅₀ (dose, which lead to a 50% inhibition) was calculated. 

1. A method of regulating meiosis comprising administering to a subject in need thereof an effective amount of the compound of formula Ic:

wherein R¹ is hydrogen, halogen, methyl, hydroxyl or oxo; R² is selected from the group consisting of hydrogen, hydroxyl, C₁-C₃ alkyl, vinyl, C₁-C₃ alkoxy and halogen; R³ is selected from the group consisting of hydrogen and optionally substituted alkoxy, acyloxy, sulphonyloxy, and phosphonyloxy; R′³ designates hydrogen or hydroxyl; R⁴ and R′⁴, which are different or identical with the proviso that they are not both hydroxyl, are selected from the group comprising hydrogen, halogen, hydroxyl and C₁-C₆ alkyl which may be substituted by halogen, hydroxyl or cyano; R⁵ is hydrogen, halogen, hydroxyl, lower alkyl, cyano, hydroxymethyl, a carbaldehyde, an oxime derived from a carbaldehyde, a carboxylic acid, a primary or secondary amine derived from a carboxylic acid or an ester with a C₁-C₆-alcohol group; R⁶ is hydrogen, hydroxyl, halogen or oxo; R⁷ is selected from the group consisting of hydrogen, hydroxyl, lower alkoxy, acyloxy, halogen and lower alkyl; R′⁷ is hydrogen, or, if R⁷ is lower alkyl, R′⁷ is hydrogen or hydroxyl; R⁸ designates together with R⁹ an additional bond between the carbon atoms at which R⁸ and R⁹ are placed; R¹¹ is selected from the group consisting of hydrogen, hydroxyl, lower alkoxy, acyloxy, halogen and lower alkyl; R′¹¹ is hydrogen or, if R¹¹ is lower alkyl, R′¹¹ is hydrogen or hydroxyl; R¹² is selected from the group consisting of hydrogen, halogen, C₁-C₄ alkyl, methylene, hydroxyl, lower alkoxy, acyloxy, oxo and a group of the general formula ═NOR³³ wherein R³³ is hydrogen or C₁-C₃ alkyl; R¹⁴ designates together with R¹⁵ an additional bond between the carbon atoms at which R¹⁴ and R¹⁵ are placed; R¹⁶ is selected from the group consisting of hydrogen, halogen, C₁-C₃ alkyl, methylene, hydroxyl, lower alkoxy, oxo and a group of the general formula ═NOR³⁴ wherein R³⁴ is hydrogen or lower alkyl; R¹⁷ is hydrogen or hydroxyl; R²⁰ is selected from the group consisting of hydrogen, lower alkyl and hydroxymethyl or R²⁰ and R′²⁰ together designate methylene or oxo; R′²⁰ is hydrogen, halogen, lower alkyl or hydroxyl; R′²² is hydrogen, hydroxyl or oxo; and R²² is cyclohexyl or cyclohexylalkyl, each optionally substituted by one or more of the following groups which substituents may be different or identical: hydroxyl, lower alkoxy, halogen, amino, cyano, carboxy, a group of the general formula —COOR³⁹, oxo, N-alkylamino or N,N-dialkylamino wherein the N-alkylamino or N,N-dialkylamino substituent optionally is substituted by carboxy, lower alkoxy or lower alkylthio; and wherein R³⁹ is lower alkyl or aralkyl.
 2. The method of claim 1, wherein the compound is (20R)-4,4-20,trimethyl-21-(cyclohexyl)-5α-pregna-8,-dien-3β-ol (20R)-4,4,20-trimethyl-21-(cyclohexyl)-5α-pregna-8,-dien-3β-ol.
 3. The method of claim 1, wherein meiosis is regulated in a mammalian germ cell.
 4. The method of claim 3, wherein the compound is administered to a germ cell located in a mammalian host.
 5. The method of claim 4, wherein the germ cell an oocyte.
 6. The method of claim 3, wherein the compound is administered to an oocyte ex vivo or in vitro.
 7. The method of claim 4, wherein the germ cell is a male germ cell.
 8. The method of claim 7, wherein the compound is administered to testicular tissue in vivo, ex vivo or in vitro. 